Update: Things mostly work, the lambda genome Bismark call was missing a space between the second file name and the –output call, which resulted in a thing that looked like filename–output which of course didn’t work. More importantly, RStudio loses it’s mind after a few samples being run, I assume from sheer volume of output, which is not good. Going to try to run this via RScript from the command line, but that will lose the record of output from here. I have the output for Bismark directed to a file, so hopefully that will preserve the important information.
This notebook is going to be an attempt to churn through all of Hollie’s RRBS Geoduck data. The rough outline of steps are as follows.
We will be aligning to two genomes, the Panopea generosa genome found here and the lambda phage genome found here
The first chunk of our code will bring us through the methylation and SNP calling in MethylExtract. This should be the most time and resource intensive portion of the analysis.
| Command Used | Arguments Used |
|---|---|
| FastQC | File Name |
| Trim Galore | –fastqc -q 20 –paired (file_R1.fq.gz file_R2.fq.gz) |
| bismark_genome_preparation | path to directory with ref. genome |
| bismark | –multicore (3) –genome (dir to genome prep) -q -1 (file_R1.fq.gz) -2 (file_R2.fq.gz) -N (1) –output_dir (dir to output) |
| MethylExtract.pl | seq=(dir to genome) inDir=(dir to bismark .sam file) minQ=(30) varFraction=(0.05) maxPval=(0.01) flagW=(99,147) flagC=(83,163) wigOut=(Y) bedOut=(Y) |
First, some general variable declarations and housekeeping stuff.
## sequence IDs from nightingales spreadsheet, this is the root of what we'll build our directory and file names from as it corresponds to individual samples.
seqID <- (c("EPI-41_S38_L005", "EPI-42_S39_L005", "EPI-43_S40_L005", "EPI-44_S41_L005", "EPI-135WG_S42_L005", "EPI-151_S2_L002", "EPI-152_S3_L002", "EPI-153_S4_L002", "EPI-154_S5_L002", "EPI-159_S6_L002", "EPI-160_S7_L002", "EPI-161_S8_L002", "EPI-162_S9_L002", "EPI-167_S10_L002", "EPI-168_S11_L002", "EPI-169_S12_L002", "EPI-170_S13_L002", "EPI-175_S14_L003", "EPI-176_S15_L003", "EPI-181_S16_L003", "EPI-182_S17_L003", "EPI-184_S18_L003", "EPI-185_S19_L003", "EPI-187_S20_L003", "EPI-188_S21_L003", "EPI-193_S22_L003", "EPI-194_S23_L003", "EPI-199_S24_L003", "EPI-200_S25_L003", "EPI-205_S26_L004", "EPI-206_S27_L004", "EPI-208_S28_L004", "EPI-209_S29_L004", "EPI-214_S30_L004", "EPI-215_S31_L004", "EPI-220_S32_L004", "EPI-221_S33_L004", "EPI-226_S34_L004","EPI-227_S35_L004", "EPI-229_S38_L004", "EPI-230_S37_L004", "EPI-103_S27_L005", "EPI-104_S28_L005", "EPI-111_S29_L005", "EPI-113_S30_L005", "EPI-119_S31_L005", "EPI-120_S32_L005", "EPI-127_S33_L005", "EPI-128_S34_L005", "EPI-135_S35_L005", "EPI-136_S36_L005", "EPI-143_S37_L005", "EPI-145_S38_L005"))
## Quick test to make sure that I can correctly cut and paste. Checks that the length of the seqID list is the same as the length of the number of unique seqIDs.
print(length(seqID))[1] 53print(length(seqID) == length(unique(seqID)))[1] TRUE## Sets our working directory where all the data is stored
setwd("~/Documents/Genewiz_hdd/")
## Generates subdirectories for each sample.
#for(i in 1:length(seqID)) {
# system(paste0("mkdir ~/Documents/Genewiz_hdd/", seqID[i], "/"))
#}
## strings correpsonding to genome files
lambda.genome <- "~/Documents/Genewiz_hdd/lambda_genome/lambda_genome.fa"
geoduck.genome <- "~/Documents/Genewiz_hdd/geo_genome/Panopea_generosa-Scaff-10k.fa"Next will be our main working chunk. I’ve decided to process each file to completion (ex. EPI-41 through MethylExtract, then EPI-42) because I feel this would be too space intensive to realistically manage with 38 samples.
## Bismark Genome Prep step. This only has to happen once, so it's run outside of the iteration step
#system(paste0("/home/shared/Bismark/bismark_genome_preparation ", geoduck.genome))
#system(paste0("/home/shared/Bismark/bismark_genome_preparation ", lambda.genome))
bismark.genome.geoduck <- "~/Documents/Genewiz_hdd/geo_genome/"
print(bismark.genome.geoduck)[1] "~/Documents/Genewiz_hdd/geo_genome/"bismark.genome.lambda <- "~/Documents/Genewiz_hdd/lambda_genome/"
print(bismark.genome.lambda)[1] "~/Documents/Genewiz_hdd/lambda_genome/"for(i in 1:length(seqID)) {
## Sets our working directory for the specific sample directory
setwd(paste0("~/Documents/Genewiz_hdd/", seqID[i], "/"))
## Prints the working directory
print(getwd())
## generates temp variables for two ends of the paired end reads
end1 <- paste0(seqID[i], "_R1_001.fastq.gz")
end2 <- paste0(seqID[i], "_R2_001.fastq.gz")
print(end1)
print(end2)
trimmedend1 <- paste0(seqID[i], "_R1_001_val_1.fq.gz")
trimmedend2 <- paste0(seqID[i], "_R2_001_val_2.fq.gz")
bismark.out <- paste0(seqID[i], "_bismark_bt2.bam")
## Untrimmed fastQC pass for R1 file
system(paste0("fastqc ~/Documents/Genewiz_hdd/", end1, " -o ~/Documents/Genewiz_hdd/untrimmed_fastqc/"))
## Untrimmed fastQC pass for R2 file
system(paste0("fastqc ~/Documents/Genewiz_hdd/", end2, " -o ~/Documents/Genewiz_hdd/untrimmed_fastqc/"))
## Trim Galore run
system(paste0("/home/shared/trimgalore/trim_galore --fastqc --fastqc_args \" -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ \" -q 20 --paired ~/Documents/Genewiz_hdd/", end1, " ~/Documents/Genewiz_hdd/", end2))
## Bismark run for lambda genome, output should appear in both the terminal window, and be directed to a text file.
system(paste0("/home/shared/Bismark/bismark --multicore 3 -N 1 -q --genome ", bismark.genome.lambda, " -1 ", trimmedend1, " -2 ", trimmedend2, "--output_dir ", getwd(), " 2>> ", seqID[i], "_bismark_lambda.txt"))
## Bismark run for geoduck genome, output should appear in both the terminal window, and be directed to a text file.
system(paste0("/home/shared/Bismark/bismark --multicore 3 -N 1 -q --genome ", bismark.genome.geoduck, " -1 ", trimmedend1, " -2 ", trimmedend2, " --output_dir ", getwd(), " 2>> ", seqID[i], "_bismark_geoduck.txt"))
## Methyl Extract run.
system(paste0("perl /home/shared/MethylExtract_1.9.1/MethylExtract.pl seq=", geoduck.genome, " inDir=", getwd(), "/ minQ=30 varFraction=0.05 maxPval=0.01 flagW=99,147 flagC=83,163 wigOut=Y bedOut=Y 2>> ", seqID[i], "_MethExt_geoduck.txt"))
## Zips up all the scaffold .sam files produced by Methyl Extract, but leaves the original .sam file and the CG and SNP summary files.
system(paste0("zip -m ", seqID[i], "-sam", getwd(), "scaffold*.sam"))
}[1] "/home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005"
[1] "EPI-41_S38_L005_R1_001.fastq.gz"
[1] "EPI-41_S38_L005_R2_001.fastq.gz"Started analysis of EPI-41_S38_L005_R1_001.fastq.gz
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Approx 95% complete for EPI-41_S38_L005_R2_001.fastq.gzAnalysis complete for EPI-41_S38_L005_R2_001.fastq.gzNo quality encoding type selected. Assuming that the data provided uses Sanger encoded Phred scores (default)
Path to Cutadapt set as: 'cutadapt' (default)1.12Cutadapt seems to be working fine (tested command 'cutadapt --version')
AUTO-DETECTING ADAPTER TYPE
===========================
Attempting to auto-detect adapter type from the first 1 million sequences of the first file (>> /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R1_001.fastq.gz <<)
Found perfect matches for the following adapter sequences:
Adapter type Count Sequence Sequences analysed Percentage
Illumina 336735 AGATCGGAAGAGC 1000000 33.67
Nextera 0 CTGTCTCTTATA 1000000 0.00
smallRNA 0 TGGAATTCTCGG 1000000 0.00
Using Illumina adapter for trimming (count: 336735). Second best hit was Nextera (count: 0)
gzip: stdout: Broken pipe
Writing report to 'EPI-41_S38_L005_R1_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R1_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-41_S38_L005_R1_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R1_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R1_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 600.78 s (23 us/read; 2.58 M reads/minute).
=== Summary ===
Total reads processed: 25,784,248
Reads with adapters: 16,123,696 (62.5%)
Reads written (passing filters): 25,784,248 (100.0%)
Total basepairs processed: 2,604,209,048 bp
Quality-trimmed: 18,982,430 bp (0.7%)
Total written (filtered): 2,166,486,166 bp (83.2%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 16123696 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 22.5%
C: 11.2%
G: 29.3%
T: 36.3%
none/other: 0.8%
Overview of removed sequences
length count expect max.err error counts
1 4261224 6446062.0 0 4261224
2 985208 1611515.5 0 985208
3 386593 402878.9 0 386593
4 267747 100719.7 0 267747
5 153542 25179.9 0 153542
6 142058 6295.0 0 142058
7 127484 1573.7 0 127484
8 141235 393.4 0 141235
9 146968 98.4 0 145699 1269
10 137799 24.6 1 130983 6816
11 148428 6.1 1 140164 8264
12 136200 1.5 1 129144 7056
13 135738 0.4 1 128519 7219
14 149617 0.4 1 140119 9498
15 147158 0.4 1 138484 8674
16 156671 0.4 1 146341 10330
17 156855 0.4 1 147063 9792
18 140692 0.4 1 133061 7631
19 153237 0.4 1 142906 10331
20 146851 0.4 1 138448 8403
21 163392 0.4 1 152027 11365
22 152561 0.4 1 143480 9081
23 145452 0.4 1 136644 8808
24 150690 0.4 1 140804 9886
25 146993 0.4 1 138057 8936
26 157169 0.4 1 146608 10561
27 152089 0.4 1 143272 8817
28 146009 0.4 1 138084 7925
29 160017 0.4 1 150511 9506
30 145418 0.4 1 138046 7372
31 159900 0.4 1 150120 9780
32 147184 0.4 1 139501 7683
33 171046 0.4 1 160373 10673
34 153175 0.4 1 145094 8081
35 155093 0.4 1 146247 8846
36 162248 0.4 1 152777 9471
37 164902 0.4 1 156272 8630
38 154829 0.4 1 145972 8857
39 167776 0.4 1 158083 9693
40 171017 0.4 1 161557 9460
41 237943 0.4 1 226993 10950
42 134399 0.4 1 128102 6297
43 70463 0.4 1 66102 4361
44 138798 0.4 1 131703 7095
45 142862 0.4 1 134968 7894
46 131743 0.4 1 124887 6856
47 147562 0.4 1 139149 8413
48 135682 0.4 1 127789 7893
49 152328 0.4 1 143084 9244
50 130308 0.4 1 123261 7047
51 135670 0.4 1 127967 7703
52 136431 0.4 1 128310 8121
53 121608 0.4 1 115016 6592
54 122994 0.4 1 115844 7150
55 134548 0.4 1 126839 7709
56 123749 0.4 1 116620 7129
57 113087 0.4 1 106349 6738
58 120087 0.4 1 113743 6344
59 116904 0.4 1 110667 6237
60 103542 0.4 1 98238 5304
61 116409 0.4 1 110389 6020
62 117850 0.4 1 111964 5886
63 105605 0.4 1 100278 5327
64 100906 0.4 1 96174 4732
65 95073 0.4 1 90841 4232
66 90350 0.4 1 86323 4027
67 92045 0.4 1 87927 4118
68 85640 0.4 1 81988 3652
69 90424 0.4 1 86699 3725
70 82750 0.4 1 79136 3614
71 85980 0.4 1 82062 3918
72 94547 0.4 1 89994 4553
73 119365 0.4 1 110854 8511
74 283657 0.4 1 273441 10216
75 165095 0.4 1 158881 6214
76 83945 0.4 1 80506 3439
77 49946 0.4 1 47787 2159
78 32346 0.4 1 30986 1360
79 20265 0.4 1 19397 868
80 14549 0.4 1 13856 693
81 10084 0.4 1 9648 436
82 7616 0.4 1 7237 379
83 5789 0.4 1 5521 268
84 4616 0.4 1 4370 246
85 3664 0.4 1 3478 186
86 2886 0.4 1 2709 177
87 2229 0.4 1 2112 117
88 1656 0.4 1 1545 111
89 1569 0.4 1 1479 90
90 1904 0.4 1 1804 100
91 2512 0.4 1 2346 166
92 3661 0.4 1 3413 248
93 8067 0.4 1 7646 421
94 22127 0.4 1 21054 1073
95 37776 0.4 1 36021 1755
96 19364 0.4 1 18276 1088
97 13594 0.4 1 12782 812
98 5497 0.4 1 5114 383
99 5436 0.4 1 5027 409
100 9228 0.4 1 8419 809
101 28701 0.4 1 25459 3242
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R1_001.fastq.gz
=============================================
25784248 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Writing report to 'EPI-41_S38_L005_R2_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R2_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-41_S38_L005_R2_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R2_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R2_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 594.26 s (23 us/read; 2.60 M reads/minute).
=== Summary ===
Total reads processed: 25,784,248
Reads with adapters: 17,635,068 (68.4%)
Reads written (passing filters): 25,784,248 (100.0%)
Total basepairs processed: 2,604,209,048 bp
Quality-trimmed: 41,974,823 bp (1.6%)
Total written (filtered): 2,163,399,405 bp (83.1%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 17635068 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 34.7%
C: 22.8%
G: 10.7%
T: 31.1%
none/other: 0.7%
Overview of removed sequences
length count expect max.err error counts
1 6957145 6446062.0 0 6957145
2 248720 1611515.5 0 248720
3 177146 402878.9 0 177146
4 147721 100719.7 0 147721
5 146383 25179.9 0 146383
6 147588 6295.0 0 147588
7 143620 1573.7 0 143620
8 145419 393.4 0 145419
9 141858 98.4 0 141272 586
10 149959 24.6 1 143538 6421
11 140286 6.1 1 132752 7534
12 148660 1.5 1 141080 7580
13 139864 0.4 1 132960 6904
14 160334 0.4 1 152041 8293
15 144459 0.4 1 137324 7135
16 148496 0.4 1 141560 6936
17 162802 0.4 1 154906 7896
18 137481 0.4 1 130751 6730
19 153879 0.4 1 146538 7341
20 149179 0.4 1 141558 7621
21 151701 0.4 1 143629 8072
22 159907 0.4 1 151511 8396
23 154426 0.4 1 146841 7585
24 178770 0.4 1 169739 9031
25 143506 0.4 1 136287 7219
26 149615 0.4 1 141325 8290
27 155343 0.4 1 145723 9620
28 172140 0.4 1 163604 8536
29 156523 0.4 1 147083 9440
30 185894 0.4 1 177226 8668
31 150699 0.4 1 142404 8295
32 171077 0.4 1 163743 7334
33 178894 0.4 1 169644 9250
34 178132 0.4 1 168112 10020
35 184751 0.4 1 177685 7066
36 161903 0.4 1 153644 8259
37 163689 0.4 1 155607 8082
38 149582 0.4 1 142404 7178
39 161959 0.4 1 153762 8197
40 158742 0.4 1 150981 7761
41 163485 0.4 1 156739 6746
42 166614 0.4 1 160560 6054
43 139347 0.4 1 132745 6602
44 151828 0.4 1 145128 6700
45 217396 0.4 1 210163 7233
46 141747 0.4 1 135687 6060
47 95032 0.4 1 90229 4803
48 155242 0.4 1 149534 5708
49 101736 0.4 1 97225 4511
50 111533 0.4 1 106532 5001
51 174495 0.4 1 168770 5725
52 96871 0.4 1 92497 4374
53 99529 0.4 1 94809 4720
54 90510 0.4 1 86254 4256
55 115293 0.4 1 110603 4690
56 111307 0.4 1 106246 5061
57 104012 0.4 1 99736 4276
58 103731 0.4 1 99505 4226
59 99979 0.4 1 95616 4363
60 99179 0.4 1 94781 4398
61 102532 0.4 1 97938 4594
62 107723 0.4 1 102747 4976
63 110480 0.4 1 105467 5013
64 110148 0.4 1 104800 5348
65 117677 0.4 1 112276 5401
66 128289 0.4 1 122114 6175
67 170411 0.4 1 159239 11172
68 446369 0.4 1 432858 13511
69 156375 0.4 1 149378 6997
70 77641 0.4 1 73323 4318
71 47002 0.4 1 43677 3325
72 36015 0.4 1 33402 2613
73 29924 0.4 1 27658 2266
74 24372 0.4 1 22530 1842
75 21368 0.4 1 19692 1676
76 18615 0.4 1 17137 1478
77 16874 0.4 1 15521 1353
78 14849 0.4 1 13623 1226
79 12362 0.4 1 11274 1088
80 10110 0.4 1 9245 865
81 8355 0.4 1 7660 695
82 6782 0.4 1 6191 591
83 5173 0.4 1 4682 491
84 4062 0.4 1 3646 416
85 3043 0.4 1 2737 306
86 2320 0.4 1 2068 252
87 1958 0.4 1 1700 258
88 1487 0.4 1 1281 206
89 1551 0.4 1 1293 258
90 1883 0.4 1 1623 260
91 2632 0.4 1 2233 399
92 3854 0.4 1 3297 557
93 7594 0.4 1 6518 1076
94 20459 0.4 1 17981 2478
95 33935 0.4 1 30160 3775
96 17519 0.4 1 15550 1969
97 12444 0.4 1 10959 1485
98 4894 0.4 1 4309 585
99 4685 0.4 1 4123 562
100 7746 0.4 1 6706 1040
101 26438 0.4 1 22711 3727
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005_R2_001.fastq.gz
=============================================
25784248 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Validate paired-end files EPI-41_S38_L005_R1_001_trimmed.fq.gz and EPI-41_S38_L005_R2_001_trimmed.fq.gz
file_1: EPI-41_S38_L005_R1_001_trimmed.fq.gz, file_2: EPI-41_S38_L005_R2_001_trimmed.fq.gz
>>>>> Now validing the length of the 2 paired-end infiles: EPI-41_S38_L005_R1_001_trimmed.fq.gz and EPI-41_S38_L005_R2_001_trimmed.fq.gz <<<<<
Writing validated paired-end read 1 reads to EPI-41_S38_L005_R1_001_val_1.fq.gz
Writing validated paired-end read 2 reads to EPI-41_S38_L005_R2_001_val_2.fq.gz
Total number of sequences analysed: 25784248
Number of sequence pairs removed because at least one read was shorter than the length cutoff (20 bp): 671841 (2.61%)
>>> Now running FastQC on the validated data EPI-41_S38_L005_R1_001_val_1.fq.gz<<<
Started analysis of EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 5% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 10% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 15% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 20% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 25% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 30% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 35% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 40% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 45% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 50% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 55% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 60% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 65% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 70% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 75% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 80% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 85% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 90% complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
Approx 95% complete for EPI-41_S38_L005_R1_001_val_1.fq.gzAnalysis complete for EPI-41_S38_L005_R1_001_val_1.fq.gz
>>> Now running FastQC on the validated data EPI-41_S38_L005_R2_001_val_2.fq.gz<<<
Started analysis of EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 5% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 10% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 15% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 20% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 25% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 30% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 35% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 40% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 45% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 50% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 55% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 60% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 65% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 70% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 75% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 80% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 85% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 90% complete for EPI-41_S38_L005_R2_001_val_2.fq.gz
Approx 95% complete for EPI-41_S38_L005_R2_001_val_2.fq.gzAnalysis complete for EPI-41_S38_L005_R2_001_val_2.fq.gzDeleting both intermediate output files EPI-41_S38_L005_R1_001_trimmed.fq.gz and EPI-41_S38_L005_R2_001_trimmed.fq.gz
====================================================================================================
gzip: stdout: Broken pipeFastQ format specified
chr J02459.1 (48502 bp)
FastQ format specified
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chr scaffold393871 (11260 bp)
chr scaffold395137 (20701 bp)
chr scaffold396627 (11762 bp)
chr scaffold398408 (10156 bp)
chr scaffold399073 (19016 bp)
chr scaffold400179 (14117 bp)
chr scaffold400577 (16728 bp)
chr scaffold402227 (11245 bp)
chr scaffold402259 (10293 bp)
chr scaffold405403 (10019 bp)
chr scaffold407741 (10885 bp)
chr scaffold410287 (12671 bp)
chr scaffold410305 (11488 bp)
chr scaffold411373 (15712 bp)
chr scaffold412266 (10423 bp)
chr scaffold413642 (10820 bp)
chr scaffold414659 (13283 bp)
chr scaffold415077 (11344 bp)
chr scaffold415997 (10225 bp)
chr scaffold416419 (17392 bp)
chr scaffold416470 (16041 bp)
chr scaffold417516 (13130 bp)
chr scaffold420597 (11379 bp)
chr scaffold420603 (10725 bp)
chr scaffold422214 (16859 bp)
chr scaffold422344 (10500 bp)
chr scaffold423561 (13732 bp)
chr scaffold424674 (10270 bp)
chr scaffold425050 (12114 bp)
chr scaffold425449 (12126 bp)
chr scaffold425469 (12533 bp)
chr scaffold428074 (12128 bp)
chr scaffold428188 (10736 bp)
chr scaffold431671 (12829 bp)
chr scaffold431819 (17424 bp)
chr scaffold431928 (10156 bp)
chr scaffold431960 (11632 bp)
chr scaffold432926 (11069 bp)
chr scaffold433332 (10628 bp)
chr scaffold434377 (13720 bp)
chr scaffold435031 (10705 bp)
chr scaffold436089 (10932 bp)
chr scaffold436358 (14110 bp)
chr scaffold436802 (11362 bp)
Number of paired-end alignments with a unique best hit: 754594
Mapping efficiency: 9.0%
Sequence pairs with no alignments under any condition: 7488167
Sequence pairs did not map uniquely: 128041
Sequence pairs which were discarded because genomic sequence could not be extracted: 117
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 379904 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 374573 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 755613
Mapping efficiency: 9.0%
Sequence pairs with no alignments under any condition: 7487342
Sequence pairs did not map uniquely: 127848
Sequence pairs which were discarded because genomic sequence could not be extracted: 117
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 380690 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 374806 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 753191
Mapping efficiency: 9.0%
Sequence pairs with no alignments under any condition: 7489212
Sequence pairs did not map uniquely: 128399
Sequence pairs which were discarded because genomic sequence could not be extracted: 154
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 379325 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 373712 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.1 EPI-41_S38_L005_R2_001_val_2.fq.gz.temp.1 EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.2 EPI-41_S38_L005_R2_001_val_2.fq.gz.temp.2 EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.3 EPI-41_S38_L005_R2_001_val_2.fq.gz.temp.3
/home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005/EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005/EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005/EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_pe.bam
Number of paired-end alignments with a unique best hit: 2263398
Mapping efficiency: 9.0%
Sequence pairs with no alignments under any condition: 22464721
Sequence pairs did not map uniquely: 384288
Sequence pairs which were discarded because genomic sequence could not be extracted: 388
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 1139919 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 1123091 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_PE_report.txt EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_PE_report.txt EPI-41_S38_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_PE_report.txt
################### Running MethylExtract v1.9 ###################
Functions summary:
Multi-fasta reference selected in ~/Documents/Genewiz_hdd/geo_genome/Panopea_generosa-Scaff-10k.fa
Watson strand FLAG: 147,99 & Crick strand FLAG: 163,83
Duplicated reads will not be deleted (default)
5' bases of the reads will not be ignored (default)
3' bases of the reads will not be ignored (default)
Pair-end overlapping parts will be included (default)
Quality values resume:
Selected Qscore: phred33-quals
Bisulfite check fraction/integer: 0.9
Bases from the 5' end to be ignored: 0
Bases from the 3' end to be ignored: 0
Minimum PHRED score for methylation and base call: 30
Minimum depth for methylation calls: 1
Minimum depth for SNV calls: 1
Alleles minimum frequency to be considered: 0.05
Maximum allowed strand bias: 0.7
SNV p-value threshold: 0.01
Activated Outputs:
Default methylation levels output: CG context
BED output format
WIG output format
SNVs VCF output format
Sorting alignments by chromosome
Reading /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005//EPI-41_S38_L005_R1_001_val_1_bismark_bt2_pe.bam
Starting main process
Merging Methylation & SNVs Outputs
Printing LOG file
# Methylation Ratios & SNVs retrieved, consult your results in /home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005/ #
# See the log file for further information #
zip error: Nothing to do! (EPI-41_S38_L005-sam/home/sean/Documents/Genewiz_hdd/EPI-41_S38_L005scaffold*.sam)
[1] "/home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005"
[1] "EPI-42_S39_L005_R1_001.fastq.gz"
[1] "EPI-42_S39_L005_R2_001.fastq.gz"Started analysis of EPI-42_S39_L005_R1_001.fastq.gz
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Approx 95% complete for EPI-42_S39_L005_R2_001.fastq.gzAnalysis complete for EPI-42_S39_L005_R2_001.fastq.gzNo quality encoding type selected. Assuming that the data provided uses Sanger encoded Phred scores (default)
Path to Cutadapt set as: 'cutadapt' (default)1.12Cutadapt seems to be working fine (tested command 'cutadapt --version')
AUTO-DETECTING ADAPTER TYPE
===========================
Attempting to auto-detect adapter type from the first 1 million sequences of the first file (>> /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R1_001.fastq.gz <<)
Found perfect matches for the following adapter sequences:
Adapter type Count Sequence Sequences analysed Percentage
Illumina 339364 AGATCGGAAGAGC 1000000 33.94
smallRNA 0 TGGAATTCTCGG 1000000 0.00
Nextera 0 CTGTCTCTTATA 1000000 0.00
Using Illumina adapter for trimming (count: 339364). Second best hit was smallRNA (count: 0)
gzip: stdout: Broken pipe
Writing report to 'EPI-42_S39_L005_R1_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R1_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-42_S39_L005_R1_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R1_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
30000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R1_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 672.23 s (22 us/read; 2.68 M reads/minute).
=== Summary ===
Total reads processed: 30,041,436
Reads with adapters: 19,010,654 (63.3%)
Reads written (passing filters): 30,041,436 (100.0%)
Total basepairs processed: 3,034,185,036 bp
Quality-trimmed: 14,121,571 bp (0.5%)
Total written (filtered): 2,544,526,180 bp (83.9%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 19010654 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 22.2%
C: 8.7%
G: 30.2%
T: 38.6%
none/other: 0.3%
Overview of removed sequences
length count expect max.err error counts
1 4839153 7510359.0 0 4839153
2 1230079 1877589.8 0 1230079
3 496881 469397.4 0 496881
4 332234 117349.4 0 332234
5 189945 29337.3 0 189945
6 178798 7334.3 0 178798
7 165879 1833.6 0 165879
8 178559 458.4 0 178559
9 182401 114.6 0 180844 1557
10 177130 28.6 1 169384 7746
11 181882 7.2 1 172935 8947
12 171867 1.8 1 163834 8033
13 171669 0.4 1 163202 8467
14 186645 0.4 1 176006 10639
15 183811 0.4 1 173927 9884
16 195217 0.4 1 183783 11434
17 190501 0.4 1 179966 10535
18 172141 0.4 1 163719 8422
19 188484 0.4 1 176755 11729
20 179359 0.4 1 170114 9245
21 205026 0.4 1 191543 13483
22 189461 0.4 1 179205 10256
23 178992 0.4 1 169034 9958
24 189011 0.4 1 177399 11612
25 182811 0.4 1 172905 9906
26 197467 0.4 1 184900 12567
27 187440 0.4 1 177488 9952
28 183510 0.4 1 174210 9300
29 200427 0.4 1 189396 11031
30 181551 0.4 1 172831 8720
31 198184 0.4 1 186977 11207
32 186477 0.4 1 177109 9368
33 203847 0.4 1 192280 11567
34 194540 0.4 1 184204 10336
35 188612 0.4 1 178870 9742
36 197121 0.4 1 187663 9458
37 196742 0.4 1 187217 9525
38 182243 0.4 1 173263 8980
39 177383 0.4 1 169016 8367
40 201535 0.4 1 189969 11566
41 273186 0.4 1 260772 12414
42 152641 0.4 1 144977 7664
43 121214 0.4 1 115030 6184
44 167704 0.4 1 159432 8272
45 173933 0.4 1 164821 9112
46 165905 0.4 1 157328 8577
47 180672 0.4 1 170644 10028
48 168762 0.4 1 159259 9503
49 179662 0.4 1 169466 10196
50 157445 0.4 1 149362 8083
51 162118 0.4 1 153422 8696
52 161449 0.4 1 152217 9232
53 151363 0.4 1 143476 7887
54 148470 0.4 1 140200 8270
55 164808 0.4 1 155888 8920
56 149600 0.4 1 141017 8583
57 139800 0.4 1 131748 8052
58 148378 0.4 1 140657 7721
59 142416 0.4 1 134970 7446
60 126969 0.4 1 120560 6409
61 139713 0.4 1 132655 7058
62 141541 0.4 1 134560 6981
63 129209 0.4 1 122589 6620
64 124623 0.4 1 118880 5743
65 117319 0.4 1 112279 5040
66 111548 0.4 1 106649 4899
67 112419 0.4 1 107493 4926
68 107295 0.4 1 102808 4487
69 113769 0.4 1 109068 4701
70 108044 0.4 1 103691 4353
71 103460 0.4 1 99169 4291
72 102749 0.4 1 98147 4602
73 126131 0.4 1 118629 7502
74 233092 0.4 1 225110 7982
75 130064 0.4 1 125412 4652
76 66306 0.4 1 63741 2565
77 40924 0.4 1 39253 1671
78 27651 0.4 1 26481 1170
79 17969 0.4 1 17114 855
80 14304 0.4 1 13686 618
81 10423 0.4 1 9985 438
82 7849 0.4 1 7528 321
83 6448 0.4 1 6188 260
84 4645 0.4 1 4448 197
85 3420 0.4 1 3264 156
86 2196 0.4 1 2076 120
87 1527 0.4 1 1441 86
88 887 0.4 1 833 54
89 713 0.4 1 660 53
90 794 0.4 1 746 48
91 909 0.4 1 849 60
92 1364 0.4 1 1260 104
93 2402 0.4 1 2259 143
94 5968 0.4 1 5617 351
95 9682 0.4 1 9197 485
96 6530 0.4 1 6147 383
97 4897 0.4 1 4556 341
98 1928 0.4 1 1781 147
99 1498 0.4 1 1333 165
100 2843 0.4 1 2468 375
101 14091 0.4 1 12101 1990
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R1_001.fastq.gz
=============================================
30041436 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Writing report to 'EPI-42_S39_L005_R2_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R2_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-42_S39_L005_R2_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R2_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
30000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R2_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 696.43 s (23 us/read; 2.59 M reads/minute).
=== Summary ===
Total reads processed: 30,041,436
Reads with adapters: 20,795,246 (69.2%)
Reads written (passing filters): 30,041,436 (100.0%)
Total basepairs processed: 3,034,185,036 bp
Quality-trimmed: 36,630,910 bp (1.2%)
Total written (filtered): 2,540,344,516 bp (83.7%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 20795246 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 35.7%
C: 23.6%
G: 9.3%
T: 31.1%
none/other: 0.3%
Overview of removed sequences
length count expect max.err error counts
1 8136675 7510359.0 0 8136675
2 251809 1877589.8 0 251809
3 230932 469397.4 0 230932
4 181015 117349.4 0 181015
5 186864 29337.3 0 186864
6 183967 7334.3 0 183967
7 177624 1833.6 0 177624
8 183343 458.4 0 183343
9 178845 114.6 0 178222 623
10 189141 28.6 1 181854 7287
11 175298 7.2 1 166402 8896
12 183586 1.8 1 174990 8596
13 174337 0.4 1 166139 8198
14 196405 0.4 1 187207 9198
15 181417 0.4 1 173210 8207
16 186101 0.4 1 177957 8144
17 196591 0.4 1 188099 8492
18 170641 0.4 1 163199 7442
19 183929 0.4 1 175883 8046
20 183745 0.4 1 175351 8394
21 188399 0.4 1 178962 9437
22 198395 0.4 1 188822 9573
23 189859 0.4 1 181499 8360
24 208299 0.4 1 198915 9384
25 181384 0.4 1 173214 8170
26 188247 0.4 1 178567 9680
27 196730 0.4 1 185317 11413
28 207628 0.4 1 198298 9330
29 197454 0.4 1 186515 10939
30 224209 0.4 1 214780 9429
31 188845 0.4 1 179382 9463
32 200318 0.4 1 192436 7882
33 215998 0.4 1 205431 10567
34 220950 0.4 1 209377 11573
35 212011 0.4 1 204273 7738
36 200582 0.4 1 190998 9584
37 200374 0.4 1 191398 8976
38 181260 0.4 1 173107 8153
39 191952 0.4 1 183040 8912
40 190909 0.4 1 182087 8822
41 194772 0.4 1 187250 7522
42 195555 0.4 1 188935 6620
43 167355 0.4 1 159901 7454
44 179529 0.4 1 172248 7281
45 245723 0.4 1 237849 7874
46 175212 0.4 1 168187 7025
47 122362 0.4 1 116622 5740
48 187347 0.4 1 180694 6653
49 127861 0.4 1 122635 5226
50 136692 0.4 1 130717 5975
51 206009 0.4 1 199505 6504
52 120762 0.4 1 115609 5153
53 125924 0.4 1 120244 5680
54 111678 0.4 1 106624 5054
55 144675 0.4 1 139144 5531
56 136968 0.4 1 131145 5823
57 128391 0.4 1 123278 5113
58 129921 0.4 1 124971 4950
59 123007 0.4 1 117818 5189
60 121787 0.4 1 116631 5156
61 124526 0.4 1 119208 5318
62 129625 0.4 1 123889 5736
63 132623 0.4 1 126952 5671
64 132349 0.4 1 126447 5902
65 139096 0.4 1 133237 5859
66 146035 0.4 1 139623 6412
67 181569 0.4 1 171576 9993
68 440815 0.4 1 429177 11638
69 157089 0.4 1 150867 6222
70 73617 0.4 1 69753 3864
71 45539 0.4 1 42636 2903
72 36979 0.4 1 34398 2581
73 32199 0.4 1 29940 2259
74 27493 0.4 1 25611 1882
75 24085 0.4 1 22355 1730
76 22046 0.4 1 20468 1578
77 20197 0.4 1 18715 1482
78 17557 0.4 1 16188 1369
79 15214 0.4 1 14054 1160
80 12541 0.4 1 11577 964
81 10613 0.4 1 9782 831
82 8104 0.4 1 7536 568
83 6403 0.4 1 5903 500
84 4716 0.4 1 4340 376
85 3171 0.4 1 2905 266
86 2007 0.4 1 1836 171
87 1400 0.4 1 1261 139
88 804 0.4 1 700 104
89 702 0.4 1 606 96
90 770 0.4 1 674 96
91 950 0.4 1 804 146
92 1352 0.4 1 1146 206
93 2171 0.4 1 1866 305
94 5407 0.4 1 4716 691
95 8719 0.4 1 7689 1030
96 5937 0.4 1 5218 719
97 4538 0.4 1 3972 566
98 1758 0.4 1 1529 229
99 1273 0.4 1 1086 187
100 2477 0.4 1 2007 470
101 13182 0.4 1 10971 2211
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005_R2_001.fastq.gz
=============================================
30041436 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Validate paired-end files EPI-42_S39_L005_R1_001_trimmed.fq.gz and EPI-42_S39_L005_R2_001_trimmed.fq.gz
file_1: EPI-42_S39_L005_R1_001_trimmed.fq.gz, file_2: EPI-42_S39_L005_R2_001_trimmed.fq.gz
>>>>> Now validing the length of the 2 paired-end infiles: EPI-42_S39_L005_R1_001_trimmed.fq.gz and EPI-42_S39_L005_R2_001_trimmed.fq.gz <<<<<
Writing validated paired-end read 1 reads to EPI-42_S39_L005_R1_001_val_1.fq.gz
Writing validated paired-end read 2 reads to EPI-42_S39_L005_R2_001_val_2.fq.gz
Total number of sequences analysed: 30041436
Number of sequence pairs removed because at least one read was shorter than the length cutoff (20 bp): 338969 (1.13%)
>>> Now running FastQC on the validated data EPI-42_S39_L005_R1_001_val_1.fq.gz<<<
Started analysis of EPI-42_S39_L005_R1_001_val_1.fq.gz
Approx 5% complete for EPI-42_S39_L005_R1_001_val_1.fq.gz
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Approx 90% complete for EPI-42_S39_L005_R1_001_val_1.fq.gz
Approx 95% complete for EPI-42_S39_L005_R1_001_val_1.fq.gzAnalysis complete for EPI-42_S39_L005_R1_001_val_1.fq.gz
>>> Now running FastQC on the validated data EPI-42_S39_L005_R2_001_val_2.fq.gz<<<
Started analysis of EPI-42_S39_L005_R2_001_val_2.fq.gz
Approx 5% complete for EPI-42_S39_L005_R2_001_val_2.fq.gz
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Approx 90% complete for EPI-42_S39_L005_R2_001_val_2.fq.gz
Approx 95% complete for EPI-42_S39_L005_R2_001_val_2.fq.gzAnalysis complete for EPI-42_S39_L005_R2_001_val_2.fq.gzDeleting both intermediate output files EPI-42_S39_L005_R1_001_trimmed.fq.gz and EPI-42_S39_L005_R2_001_trimmed.fq.gz
====================================================================================================
gzip: stdout: Broken pipeFastQ format specified
chr J02459.1 (48502 bp)
FastQ format specified
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chr scaffold414659 (13283 bp)
chr scaffold415077 (11344 bp)
chr scaffold415997 (10225 bp)
chr scaffold416419 (17392 bp)
chr scaffold416470 (16041 bp)
chr scaffold417516 (13130 bp)
chr scaffold420597 (11379 bp)
chr scaffold420603 (10725 bp)
chr scaffold422214 (16859 bp)
chr scaffold422344 (10500 bp)
chr scaffold423561 (13732 bp)
chr scaffold424674 (10270 bp)
chr scaffold425050 (12114 bp)
chr scaffold425449 (12126 bp)
chr scaffold425469 (12533 bp)
chr scaffold428074 (12128 bp)
chr scaffold428188 (10736 bp)
chr scaffold431671 (12829 bp)
chr scaffold431819 (17424 bp)
chr scaffold431928 (10156 bp)
chr scaffold431960 (11632 bp)
chr scaffold432926 (11069 bp)
chr scaffold433332 (10628 bp)
chr scaffold434377 (13720 bp)
chr scaffold435031 (10705 bp)
chr scaffold436089 (10932 bp)
chr scaffold436358 (14110 bp)
chr scaffold436802 (11362 bp)
Number of paired-end alignments with a unique best hit: 1036578
Mapping efficiency: 10.5%
Sequence pairs with no alignments under any condition: 8682215
Sequence pairs did not map uniquely: 182029
Sequence pairs which were discarded because genomic sequence could not be extracted: 188
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 522196 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 514194 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 1039043
Mapping efficiency: 10.5%
Sequence pairs with no alignments under any condition: 8680201
Sequence pairs did not map uniquely: 181579
Sequence pairs which were discarded because genomic sequence could not be extracted: 196
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 522747 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 516100 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 1036580
Mapping efficiency: 10.5%
Sequence pairs with no alignments under any condition: 8682398
Sequence pairs did not map uniquely: 181844
Sequence pairs which were discarded because genomic sequence could not be extracted: 186
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 522318 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 514076 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.1 EPI-42_S39_L005_R2_001_val_2.fq.gz.temp.1 EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.2 EPI-42_S39_L005_R2_001_val_2.fq.gz.temp.2 EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.3 EPI-42_S39_L005_R2_001_val_2.fq.gz.temp.3
/home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005/EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005/EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005/EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_pe.bam
Number of paired-end alignments with a unique best hit: 3112201
Mapping efficiency: 10.5%
Sequence pairs with no alignments under any condition: 26044814
Sequence pairs did not map uniquely: 545452
Sequence pairs which were discarded because genomic sequence could not be extracted: 570
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 1567261 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 1544370 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_PE_report.txt EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_PE_report.txt EPI-42_S39_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_PE_report.txt
################### Running MethylExtract v1.9 ###################
Functions summary:
Multi-fasta reference selected in ~/Documents/Genewiz_hdd/geo_genome/Panopea_generosa-Scaff-10k.fa
Watson strand FLAG: 99,147 & Crick strand FLAG: 163,83
Duplicated reads will not be deleted (default)
5' bases of the reads will not be ignored (default)
3' bases of the reads will not be ignored (default)
Pair-end overlapping parts will be included (default)
Quality values resume:
Selected Qscore: phred33-quals
Bisulfite check fraction/integer: 0.9
Bases from the 5' end to be ignored: 0
Bases from the 3' end to be ignored: 0
Minimum PHRED score for methylation and base call: 30
Minimum depth for methylation calls: 1
Minimum depth for SNV calls: 1
Alleles minimum frequency to be considered: 0.05
Maximum allowed strand bias: 0.7
SNV p-value threshold: 0.01
Activated Outputs:
Default methylation levels output: CG context
BED output format
WIG output format
SNVs VCF output format
Sorting alignments by chromosome
Reading /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005//EPI-42_S39_L005_R1_001_val_1_bismark_bt2_pe.bam
Starting main process
Merging Methylation & SNVs Outputs
Printing LOG file
# Methylation Ratios & SNVs retrieved, consult your results in /home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005/ #
# See the log file for further information #
zip error: Nothing to do! (EPI-42_S39_L005-sam/home/sean/Documents/Genewiz_hdd/EPI-42_S39_L005scaffold*.sam)
[1] "/home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005"
[1] "EPI-43_S40_L005_R1_001.fastq.gz"
[1] "EPI-43_S40_L005_R2_001.fastq.gz"Started analysis of EPI-43_S40_L005_R1_001.fastq.gz
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Approx 95% complete for EPI-43_S40_L005_R2_001.fastq.gzAnalysis complete for EPI-43_S40_L005_R2_001.fastq.gzNo quality encoding type selected. Assuming that the data provided uses Sanger encoded Phred scores (default)
Path to Cutadapt set as: 'cutadapt' (default)1.12Cutadapt seems to be working fine (tested command 'cutadapt --version')
AUTO-DETECTING ADAPTER TYPE
===========================
Attempting to auto-detect adapter type from the first 1 million sequences of the first file (>> /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R1_001.fastq.gz <<)
Found perfect matches for the following adapter sequences:
Adapter type Count Sequence Sequences analysed Percentage
Illumina 458788 AGATCGGAAGAGC 1000000 45.88
smallRNA 0 TGGAATTCTCGG 1000000 0.00
Nextera 0 CTGTCTCTTATA 1000000 0.00
Using Illumina adapter for trimming (count: 458788). Second best hit was smallRNA (count: 0)
gzip: stdout: Broken pipe
Writing report to 'EPI-43_S40_L005_R1_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R1_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-43_S40_L005_R1_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R1_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R1_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 668.47 s (23 us/read; 2.64 M reads/minute).
=== Summary ===
Total reads processed: 29,360,787
Reads with adapters: 20,620,662 (70.2%)
Reads written (passing filters): 29,360,787 (100.0%)
Total basepairs processed: 2,965,439,487 bp
Quality-trimmed: 107,978,849 bp (3.6%)
Total written (filtered): 1,998,784,073 bp (67.4%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 20620662 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 20.0%
C: 26.5%
G: 21.6%
T: 30.4%
none/other: 1.5%
Overview of removed sequences
length count expect max.err error counts
1 3895309 7340196.8 0 3895309
2 911786 1835049.2 0 911786
3 364394 458762.3 0 364394
4 245445 114690.6 0 245445
5 137427 28672.6 0 137427
6 130590 7168.2 0 130590
7 118550 1792.0 0 118550
8 128937 448.0 0 128937
9 134687 112.0 0 133531 1156
10 128672 28.0 1 122607 6065
11 133534 7.0 1 126414 7120
12 124322 1.8 1 118184 6138
13 126893 0.4 1 120474 6419
14 137165 0.4 1 128979 8186
15 134894 0.4 1 127260 7634
16 141806 0.4 1 133073 8733
17 143893 0.4 1 135223 8670
18 126217 0.4 1 119575 6642
19 140931 0.4 1 131535 9396
20 132861 0.4 1 125609 7252
21 150744 0.4 1 140261 10483
22 138844 0.4 1 130921 7923
23 132812 0.4 1 125079 7733
24 140093 0.4 1 130865 9228
25 136841 0.4 1 128749 8092
26 145127 0.4 1 135527 9600
27 140613 0.4 1 132746 7867
28 135087 0.4 1 127863 7224
29 147522 0.4 1 138906 8616
30 137145 0.4 1 130148 6997
31 149534 0.4 1 140339 9195
32 140476 0.4 1 133050 7426
33 143670 0.4 1 135704 7966
34 141834 0.4 1 134174 7660
35 150971 0.4 1 142012 8959
36 139448 0.4 1 132506 6942
37 147735 0.4 1 139832 7903
38 144478 0.4 1 136801 7677
39 139182 0.4 1 132134 7048
40 159075 0.4 1 149579 9496
41 211374 0.4 1 200982 10392
42 124477 0.4 1 118619 5858
43 69394 0.4 1 65348 4046
44 126800 0.4 1 120451 6349
45 129981 0.4 1 122803 7178
46 124010 0.4 1 117422 6588
47 137187 0.4 1 129398 7789
48 126428 0.4 1 119319 7109
49 136024 0.4 1 127949 8075
50 117632 0.4 1 111322 6310
51 123096 0.4 1 116072 7024
52 122457 0.4 1 115148 7309
53 112318 0.4 1 106214 6104
54 109531 0.4 1 103176 6355
55 122692 0.4 1 115625 7067
56 111022 0.4 1 104673 6349
57 99646 0.4 1 93864 5782
58 109067 0.4 1 103391 5676
59 103507 0.4 1 98170 5337
60 90490 0.4 1 85927 4563
61 103419 0.4 1 98033 5386
62 104191 0.4 1 98953 5238
63 89905 0.4 1 85415 4490
64 87225 0.4 1 83121 4104
65 80646 0.4 1 76974 3672
66 77685 0.4 1 74120 3565
67 79888 0.4 1 76267 3621
68 80567 0.4 1 76905 3662
69 91569 0.4 1 87446 4123
70 94166 0.4 1 89765 4401
71 113625 0.4 1 108163 5462
72 160639 0.4 1 150732 9907
73 445515 0.4 1 410099 35416
74 1830536 0.4 1 1771323 59213
75 1038058 0.4 1 999743 38315
76 575820 0.4 1 553705 22115
77 318777 0.4 1 306268 12509
78 184889 0.4 1 177546 7343
79 100125 0.4 1 95908 4217
80 66140 0.4 1 63314 2826
81 41376 0.4 1 39548 1828
82 28595 0.4 1 27253 1342
83 22547 0.4 1 21372 1175
84 19040 0.4 1 18051 989
85 17791 0.4 1 16841 950
86 17093 0.4 1 16158 935
87 15693 0.4 1 14835 858
88 14265 0.4 1 13497 768
89 15479 0.4 1 14615 864
90 20502 0.4 1 19472 1030
91 31226 0.4 1 29708 1518
92 52453 0.4 1 49824 2629
93 134451 0.4 1 128032 6419
94 399545 0.4 1 382017 17528
95 644966 0.4 1 616839 28127
96 263844 0.4 1 251156 12688
97 137147 0.4 1 129773 7374
98 50714 0.4 1 47862 2852
99 47408 0.4 1 44737 2671
100 44576 0.4 1 41826 2750
101 71889 0.4 1 66438 5451
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R1_001.fastq.gz
=============================================
29360787 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Writing report to 'EPI-43_S40_L005_R2_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R2_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-43_S40_L005_R2_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R2_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R2_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 673.00 s (23 us/read; 2.62 M reads/minute).
=== Summary ===
Total reads processed: 29,360,787
Reads with adapters: 21,882,728 (74.5%)
Reads written (passing filters): 29,360,787 (100.0%)
Total basepairs processed: 2,965,439,487 bp
Quality-trimmed: 164,608,425 bp (5.6%)
Total written (filtered): 2,006,745,702 bp (67.7%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 21882728 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 29.3%
C: 18.6%
G: 19.4%
T: 31.2%
none/other: 1.4%
Overview of removed sequences
length count expect max.err error counts
1 6425203 7340196.8 0 6425203
2 222611 1835049.2 0 222611
3 166260 458762.3 0 166260
4 134732 114690.6 0 134732
5 134196 28672.6 0 134196
6 135011 7168.2 0 135011
7 131257 1792.0 0 131257
8 133293 448.0 0 133293
9 133072 112.0 0 132508 564
10 139083 28.0 1 133494 5589
11 129206 7.0 1 122603 6603
12 135888 1.8 1 129075 6813
13 130583 0.4 1 124481 6102
14 147134 0.4 1 139732 7402
15 133833 0.4 1 127812 6021
16 137078 0.4 1 131058 6020
17 149334 0.4 1 142774 6560
18 125534 0.4 1 120077 5457
19 139649 0.4 1 133483 6166
20 136761 0.4 1 130268 6493
21 140175 0.4 1 133100 7075
22 147271 0.4 1 139958 7313
23 143543 0.4 1 136929 6614
24 163549 0.4 1 155622 7927
25 134196 0.4 1 127862 6334
26 141843 0.4 1 134155 7688
27 147300 0.4 1 138248 9052
28 160950 0.4 1 153248 7702
29 147584 0.4 1 139327 8257
30 178101 0.4 1 170092 8009
31 145894 0.4 1 137843 8051
32 165208 0.4 1 158020 7188
33 172896 0.4 1 163686 9210
34 172809 0.4 1 162944 9865
35 176380 0.4 1 169628 6752
36 153064 0.4 1 145462 7602
37 154832 0.4 1 147524 7308
38 141291 0.4 1 134489 6802
39 152931 0.4 1 145358 7573
40 149737 0.4 1 142488 7249
41 155137 0.4 1 148885 6252
42 164082 0.4 1 158039 6043
43 131622 0.4 1 125602 6020
44 148796 0.4 1 142325 6471
45 244817 0.4 1 236533 8284
46 139728 0.4 1 133899 5829
47 93015 0.4 1 88475 4540
48 157918 0.4 1 152203 5715
49 96554 0.4 1 92481 4073
50 105268 0.4 1 100746 4522
51 172026 0.4 1 166528 5498
52 91291 0.4 1 87197 4094
53 94297 0.4 1 90063 4234
54 85172 0.4 1 81377 3795
55 109874 0.4 1 105657 4217
56 103483 0.4 1 99086 4397
57 97169 0.4 1 93389 3780
58 99638 0.4 1 95674 3964
59 92605 0.4 1 88599 4006
60 92416 0.4 1 88381 4035
61 97686 0.4 1 93112 4574
62 106561 0.4 1 101399 5162
63 116927 0.4 1 110869 6058
64 131591 0.4 1 124216 7375
65 167842 0.4 1 158219 9623
66 239067 0.4 1 224094 14973
67 492650 0.4 1 450060 42590
68 2022410 0.4 1 1948510 73900
69 752699 0.4 1 718139 34560
70 412515 0.4 1 392341 20174
71 210792 0.4 1 198600 12192
72 136959 0.4 1 128621 8338
73 89374 0.4 1 83274 6100
74 67577 0.4 1 62662 4915
75 53321 0.4 1 49250 4071
76 43762 0.4 1 40200 3562
77 38046 0.4 1 34750 3296
78 33166 0.4 1 30233 2933
79 28673 0.4 1 25998 2675
80 25245 0.4 1 22771 2474
81 22568 0.4 1 20288 2280
82 20201 0.4 1 18094 2107
83 18379 0.4 1 16373 2006
84 16756 0.4 1 14770 1986
85 15604 0.4 1 13663 1941
86 15098 0.4 1 13174 1924
87 15061 0.4 1 13131 1930
88 16275 0.4 1 14107 2168
89 18383 0.4 1 15987 2396
90 24307 0.4 1 21266 3041
91 35914 0.4 1 31520 4394
92 56419 0.4 1 49589 6830
93 126394 0.4 1 112535 13859
94 368652 0.4 1 332833 35819
95 594507 0.4 1 539514 54993
96 245129 0.4 1 222974 22155
97 126946 0.4 1 115180 11766
98 45559 0.4 1 41427 4132
99 42030 0.4 1 38026 4004
100 38363 0.4 1 34679 3684
101 69140 0.4 1 62160 6980
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005_R2_001.fastq.gz
=============================================
29360787 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Validate paired-end files EPI-43_S40_L005_R1_001_trimmed.fq.gz and EPI-43_S40_L005_R2_001_trimmed.fq.gz
file_1: EPI-43_S40_L005_R1_001_trimmed.fq.gz, file_2: EPI-43_S40_L005_R2_001_trimmed.fq.gz
>>>>> Now validing the length of the 2 paired-end infiles: EPI-43_S40_L005_R1_001_trimmed.fq.gz and EPI-43_S40_L005_R2_001_trimmed.fq.gz <<<<<
Writing validated paired-end read 1 reads to EPI-43_S40_L005_R1_001_val_1.fq.gz
Writing validated paired-end read 2 reads to EPI-43_S40_L005_R2_001_val_2.fq.gz
Total number of sequences analysed: 29360787
Number of sequence pairs removed because at least one read was shorter than the length cutoff (20 bp): 6752613 (23.00%)
>>> Now running FastQC on the validated data EPI-43_S40_L005_R1_001_val_1.fq.gz<<<
Started analysis of EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 5% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 10% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 15% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 20% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 25% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 30% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 35% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 40% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 45% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 50% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 55% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 60% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 65% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 70% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 75% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 80% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 85% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 90% complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
Approx 95% complete for EPI-43_S40_L005_R1_001_val_1.fq.gzAnalysis complete for EPI-43_S40_L005_R1_001_val_1.fq.gz
>>> Now running FastQC on the validated data EPI-43_S40_L005_R2_001_val_2.fq.gz<<<
Started analysis of EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 5% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 10% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 15% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 20% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 25% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 30% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 35% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 40% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 45% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 50% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 55% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 60% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 65% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 70% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 75% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 80% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 85% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 90% complete for EPI-43_S40_L005_R2_001_val_2.fq.gz
Approx 95% complete for EPI-43_S40_L005_R2_001_val_2.fq.gzAnalysis complete for EPI-43_S40_L005_R2_001_val_2.fq.gzDeleting both intermediate output files EPI-43_S40_L005_R1_001_trimmed.fq.gz and EPI-43_S40_L005_R2_001_trimmed.fq.gz
====================================================================================================
gzip: stdout: Broken pipeFastQ format specified
chr J02459.1 (48502 bp)
FastQ format specified
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chr scaffold422344 (10500 bp)
chr scaffold423561 (13732 bp)
chr scaffold424674 (10270 bp)
chr scaffold425050 (12114 bp)
chr scaffold425449 (12126 bp)
chr scaffold425469 (12533 bp)
chr scaffold428074 (12128 bp)
chr scaffold428188 (10736 bp)
chr scaffold431671 (12829 bp)
chr scaffold431819 (17424 bp)
chr scaffold431928 (10156 bp)
chr scaffold431960 (11632 bp)
chr scaffold432926 (11069 bp)
chr scaffold433332 (10628 bp)
chr scaffold434377 (13720 bp)
chr scaffold435031 (10705 bp)
chr scaffold436089 (10932 bp)
chr scaffold436358 (14110 bp)
chr scaffold436802 (11362 bp)
Number of paired-end alignments with a unique best hit: 795213
Mapping efficiency: 10.6%
Sequence pairs with no alignments under any condition: 6614280
Sequence pairs did not map uniquely: 126565
Sequence pairs which were discarded because genomic sequence could not be extracted: 133
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 399135 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 395945 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 795345
Mapping efficiency: 10.6%
Sequence pairs with no alignments under any condition: 6614426
Sequence pairs did not map uniquely: 126287
Sequence pairs which were discarded because genomic sequence could not be extracted: 123
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 400018 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 395204 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 795769
Mapping efficiency: 10.6%
Sequence pairs with no alignments under any condition: 6613224
Sequence pairs did not map uniquely: 127065
Sequence pairs which were discarded because genomic sequence could not be extracted: 126
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 399824 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 395819 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.1 EPI-43_S40_L005_R2_001_val_2.fq.gz.temp.1 EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.2 EPI-43_S40_L005_R2_001_val_2.fq.gz.temp.2 EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.3 EPI-43_S40_L005_R2_001_val_2.fq.gz.temp.3
/home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005/EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005/EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005/EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_pe.bam
Number of paired-end alignments with a unique best hit: 2386327
Mapping efficiency: 10.6%
Sequence pairs with no alignments under any condition: 19841930
Sequence pairs did not map uniquely: 379917
Sequence pairs which were discarded because genomic sequence could not be extracted: 382
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 1198977 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 1186968 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_PE_report.txt EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_PE_report.txt EPI-43_S40_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_PE_report.txt
################### Running MethylExtract v1.9 ###################
Functions summary:
Multi-fasta reference selected in ~/Documents/Genewiz_hdd/geo_genome/Panopea_generosa-Scaff-10k.fa
Watson strand FLAG: 147,99 & Crick strand FLAG: 83,163
Duplicated reads will not be deleted (default)
5' bases of the reads will not be ignored (default)
3' bases of the reads will not be ignored (default)
Pair-end overlapping parts will be included (default)
Quality values resume:
Selected Qscore: phred33-quals
Bisulfite check fraction/integer: 0.9
Bases from the 5' end to be ignored: 0
Bases from the 3' end to be ignored: 0
Minimum PHRED score for methylation and base call: 30
Minimum depth for methylation calls: 1
Minimum depth for SNV calls: 1
Alleles minimum frequency to be considered: 0.05
Maximum allowed strand bias: 0.7
SNV p-value threshold: 0.01
Activated Outputs:
Default methylation levels output: CG context
BED output format
WIG output format
SNVs VCF output format
Sorting alignments by chromosome
Reading /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005//EPI-43_S40_L005_R1_001_val_1_bismark_bt2_pe.bam
Starting main process
Merging Methylation & SNVs Outputs
Printing LOG file
# Methylation Ratios & SNVs retrieved, consult your results in /home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005/ #
# See the log file for further information #
zip error: Nothing to do! (EPI-43_S40_L005-sam/home/sean/Documents/Genewiz_hdd/EPI-43_S40_L005scaffold*.sam)
[1] "/home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005"
[1] "EPI-44_S41_L005_R1_001.fastq.gz"
[1] "EPI-44_S41_L005_R2_001.fastq.gz"Started analysis of EPI-44_S41_L005_R1_001.fastq.gz
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Approx 95% complete for EPI-44_S41_L005_R2_001.fastq.gzAnalysis complete for EPI-44_S41_L005_R2_001.fastq.gzNo quality encoding type selected. Assuming that the data provided uses Sanger encoded Phred scores (default)
Path to Cutadapt set as: 'cutadapt' (default)1.12Cutadapt seems to be working fine (tested command 'cutadapt --version')
AUTO-DETECTING ADAPTER TYPE
===========================
Attempting to auto-detect adapter type from the first 1 million sequences of the first file (>> /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R1_001.fastq.gz <<)
Found perfect matches for the following adapter sequences:
Adapter type Count Sequence Sequences analysed Percentage
Illumina 733749 AGATCGGAAGAGC 1000000 73.37
smallRNA 0 TGGAATTCTCGG 1000000 0.00
Nextera 0 CTGTCTCTTATA 1000000 0.00
Using Illumina adapter for trimming (count: 733749). Second best hit was smallRNA (count: 0)
gzip: stdout: Broken pipe
Writing report to 'EPI-44_S41_L005_R1_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R1_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-44_S41_L005_R1_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R1_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
30000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R1_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 841.98 s (22 us/read; 2.77 M reads/minute).
=== Summary ===
Total reads processed: 38,920,722
Reads with adapters: 33,753,969 (86.7%)
Reads written (passing filters): 38,920,722 (100.0%)
Total basepairs processed: 3,930,992,922 bp
Quality-trimmed: 369,582,117 bp (9.4%)
Total written (filtered): 1,308,776,257 bp (33.3%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 33753969 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 15.6%
C: 49.5%
G: 11.2%
T: 19.2%
none/other: 4.4%
Overview of removed sequences
length count expect max.err error counts
1 2342573 9730180.5 0 2342573
2 554938 2432545.1 0 554938
3 221265 608136.3 0 221265
4 145862 152034.1 0 145862
5 79262 38008.5 0 79262
6 74365 9502.1 0 74365
7 68535 2375.5 0 68535
8 73161 593.9 0 73161
9 75634 148.5 0 74906 728
10 73838 37.1 1 70554 3284
11 76989 9.3 1 73011 3978
12 71519 2.3 1 68135 3384
13 72547 0.6 1 68937 3610
14 78117 0.6 1 73480 4637
15 77734 0.6 1 73487 4247
16 82661 0.6 1 77734 4927
17 81476 0.6 1 76679 4797
18 73276 0.6 1 69726 3550
19 81430 0.6 1 76202 5228
20 75957 0.6 1 71914 4043
21 88967 0.6 1 82938 6029
22 80446 0.6 1 76041 4405
23 76909 0.6 1 72469 4440
24 82026 0.6 1 76846 5180
25 79151 0.6 1 74633 4518
26 86459 0.6 1 80705 5754
27 81877 0.6 1 77304 4573
28 79192 0.6 1 75012 4180
29 86917 0.6 1 81848 5069
30 79601 0.6 1 75563 4038
31 86906 0.6 1 81731 5175
32 84384 0.6 1 79799 4585
33 85562 0.6 1 80892 4670
34 82994 0.6 1 78724 4270
35 80689 0.6 1 76590 4099
36 84355 0.6 1 79712 4643
37 93276 0.6 1 88115 5161
38 89826 0.6 1 84466 5360
39 86579 0.6 1 82054 4525
40 90843 0.6 1 85488 5355
41 126158 0.6 1 119845 6313
42 78660 0.6 1 74942 3718
43 45571 0.6 1 42883 2688
44 79131 0.6 1 75220 3911
45 81433 0.6 1 76954 4479
46 76799 0.6 1 72769 4030
47 86512 0.6 1 81461 5051
48 81813 0.6 1 77114 4699
49 87931 0.6 1 82571 5360
50 77386 0.6 1 73221 4165
51 80366 0.6 1 75725 4641
52 83103 0.6 1 78038 5065
53 75421 0.6 1 71345 4076
54 74440 0.6 1 70157 4283
55 83961 0.6 1 79105 4856
56 77031 0.6 1 72585 4446
57 72414 0.6 1 67939 4475
58 80952 0.6 1 76591 4361
59 77285 0.6 1 72993 4292
60 69052 0.6 1 65262 3790
61 84855 0.6 1 80251 4604
62 86341 0.6 1 81912 4429
63 78019 0.6 1 73748 4271
64 76757 0.6 1 73092 3665
65 71577 0.6 1 68252 3325
66 76047 0.6 1 72227 3820
67 88133 0.6 1 83699 4434
68 104595 0.6 1 99082 5513
69 142040 0.6 1 134938 7102
70 176038 0.6 1 166617 9421
71 273510 0.6 1 259119 14391
72 428585 0.6 1 401025 27560
73 1192617 0.6 1 1088803 103814
74 5405423 0.6 1 5221238 184185
75 3785488 0.6 1 3647387 138101
76 2430968 0.6 1 2340247 90721
77 1515538 0.6 1 1458921 56617
78 918766 0.6 1 884927 33839
79 500382 0.6 1 480379 20003
80 321065 0.6 1 308148 12917
81 196010 0.6 1 187564 8446
82 131075 0.6 1 124825 6250
83 102704 0.6 1 97686 5018
84 85341 0.6 1 80883 4458
85 79754 0.6 1 75579 4175
86 76363 0.6 1 72485 3878
87 68380 0.6 1 64669 3711
88 61474 0.6 1 58069 3405
89 61699 0.6 1 58308 3391
90 77739 0.6 1 73602 4137
91 107932 0.6 1 102340 5592
92 182417 0.6 1 173166 9251
93 475162 0.6 1 452932 22230
94 1427275 0.6 1 1364226 63049
95 2423675 0.6 1 2318373 105302
96 1068813 0.6 1 1018256 50557
97 608393 0.6 1 576295 32098
98 226087 0.6 1 213399 12688
99 217668 0.6 1 205620 12048
100 211381 0.6 1 198861 12520
101 344366 0.6 1 320919 23447
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R1_001.fastq.gz
=============================================
38920722 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Writing report to 'EPI-44_S41_L005_R2_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R2_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-44_S41_L005_R2_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R2_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
30000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R2_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 851.68 s (22 us/read; 2.74 M reads/minute).
=== Summary ===
Total reads processed: 38,920,722
Reads with adapters: 34,034,770 (87.4%)
Reads written (passing filters): 38,920,722 (100.0%)
Total basepairs processed: 3,930,992,922 bp
Quality-trimmed: 545,664,145 bp (13.9%)
Total written (filtered): 1,354,648,320 bp (34.5%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 34034770 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 17.6%
C: 14.2%
G: 34.1%
T: 29.7%
none/other: 4.3%
Overview of removed sequences
length count expect max.err error counts
1 4074995 9730180.5 0 4074995
2 150796 2432545.1 0 150796
3 110881 608136.3 0 110881
4 83450 152034.1 0 83450
5 83082 38008.5 0 83082
6 79646 9502.1 0 79646
7 78217 2375.5 0 78217
8 77680 593.9 0 77680
9 77380 148.5 0 76978 402
10 83506 37.1 1 78700 4806
11 78901 9.3 1 73026 5875
12 81137 2.3 1 75923 5214
13 77179 0.6 1 72351 4828
14 87387 0.6 1 81923 5464
15 79803 0.6 1 75067 4736
16 81714 0.6 1 76947 4767
17 86724 0.6 1 81883 4841
18 75824 0.6 1 71493 4331
19 83650 0.6 1 78954 4696
20 82847 0.6 1 77419 5428
21 87512 0.6 1 80925 6587
22 91237 0.6 1 85061 6176
23 88534 0.6 1 82881 5653
24 103701 0.6 1 96447 7254
25 83744 0.6 1 78383 5361
26 92879 0.6 1 85669 7210
27 100760 0.6 1 91537 9223
28 108001 0.6 1 100271 7730
29 102558 0.6 1 93709 8849
30 128198 0.6 1 119349 8849
31 112817 0.6 1 103247 9570
32 132316 0.6 1 123828 8488
33 144779 0.6 1 133305 11474
34 142120 0.6 1 130171 11949
35 140092 0.6 1 132254 7838
36 118156 0.6 1 109604 8552
37 120242 0.6 1 112078 8164
38 105370 0.6 1 97746 7624
39 121778 0.6 1 112914 8864
40 116337 0.6 1 107603 8734
41 122141 0.6 1 114885 7256
42 140778 0.6 1 133375 7403
43 99358 0.6 1 92625 6733
44 129719 0.6 1 121449 8270
45 313162 0.6 1 300483 12679
46 117442 0.6 1 110762 6680
47 72394 0.6 1 67542 4852
48 143808 0.6 1 136937 6871
49 72555 0.6 1 68459 4096
50 78978 0.6 1 74292 4686
51 155306 0.6 1 148690 6616
52 75507 0.6 1 71093 4414
53 73543 0.6 1 69310 4233
54 66577 0.6 1 62828 3749
55 86159 0.6 1 81639 4520
56 82189 0.6 1 77659 4530
57 81549 0.6 1 77255 4294
58 90865 0.6 1 85746 5119
59 82082 0.6 1 77095 4987
60 86598 0.6 1 81329 5269
61 101786 0.6 1 94987 6799
62 129752 0.6 1 120695 9057
63 170174 0.6 1 157607 12567
64 233751 0.6 1 216283 17468
65 361445 0.6 1 335467 25978
66 624351 0.6 1 578601 45750
67 1524642 0.6 1 1379473 145169
68 6886361 0.6 1 6624061 262300
69 2630499 0.6 1 2507064 123435
70 1469283 0.6 1 1397120 72163
71 739467 0.6 1 697053 42414
72 467157 0.6 1 438221 28936
73 294597 0.6 1 274344 20253
74 220175 0.6 1 203651 16524
75 170816 0.6 1 156939 13877
76 140106 0.6 1 128112 11994
77 121681 0.6 1 110866 10815
78 105286 0.6 1 95457 9829
79 93535 0.6 1 84288 9247
80 84606 0.6 1 76002 8604
81 76709 0.6 1 68755 7954
82 69613 0.6 1 62076 7537
83 65399 0.6 1 57958 7441
84 60054 0.6 1 52742 7312
85 56331 0.6 1 49321 7010
86 55226 0.6 1 48203 7023
87 55866 0.6 1 48622 7244
88 59152 0.6 1 51326 7826
89 66606 0.6 1 57713 8893
90 84135 0.6 1 73225 10910
91 119826 0.6 1 104730 15096
92 186555 0.6 1 163511 23044
93 427439 0.6 1 378694 48745
94 1285470 0.6 1 1156506 128964
95 2220079 0.6 1 2011361 208718
96 981495 0.6 1 892515 88980
97 564208 0.6 1 511482 52726
98 200736 0.6 1 181832 18904
99 192347 0.6 1 173735 18612
100 180192 0.6 1 162571 17621
101 331222 0.6 1 298161 33061
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005_R2_001.fastq.gz
=============================================
38920722 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Validate paired-end files EPI-44_S41_L005_R1_001_trimmed.fq.gz and EPI-44_S41_L005_R2_001_trimmed.fq.gz
file_1: EPI-44_S41_L005_R1_001_trimmed.fq.gz, file_2: EPI-44_S41_L005_R2_001_trimmed.fq.gz
>>>>> Now validing the length of the 2 paired-end infiles: EPI-44_S41_L005_R1_001_trimmed.fq.gz and EPI-44_S41_L005_R2_001_trimmed.fq.gz <<<<<
Writing validated paired-end read 1 reads to EPI-44_S41_L005_R1_001_val_1.fq.gz
Writing validated paired-end read 2 reads to EPI-44_S41_L005_R2_001_val_2.fq.gz
Total number of sequences analysed: 38920722
Number of sequence pairs removed because at least one read was shorter than the length cutoff (20 bp): 25113397 (64.52%)
>>> Now running FastQC on the validated data EPI-44_S41_L005_R1_001_val_1.fq.gz<<<
Started analysis of EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 5% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 10% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 15% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 20% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 25% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 30% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 35% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 40% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 45% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 50% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 55% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 60% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 65% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 70% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 75% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 80% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 85% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 90% complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
Approx 95% complete for EPI-44_S41_L005_R1_001_val_1.fq.gzAnalysis complete for EPI-44_S41_L005_R1_001_val_1.fq.gz
>>> Now running FastQC on the validated data EPI-44_S41_L005_R2_001_val_2.fq.gz<<<
Started analysis of EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 5% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 10% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 15% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 20% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 25% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 30% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 35% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 40% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 45% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 50% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 55% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 60% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 65% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 70% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 75% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 80% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 85% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 90% complete for EPI-44_S41_L005_R2_001_val_2.fq.gz
Approx 95% complete for EPI-44_S41_L005_R2_001_val_2.fq.gzAnalysis complete for EPI-44_S41_L005_R2_001_val_2.fq.gzDeleting both intermediate output files EPI-44_S41_L005_R1_001_trimmed.fq.gz and EPI-44_S41_L005_R2_001_trimmed.fq.gz
====================================================================================================
gzip: stdout: Broken pipeFastQ format specified
chr J02459.1 (48502 bp)
FastQ format specified
chr scaffold3 (23590 bp)
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chr scaffold422344 (10500 bp)
chr scaffold423561 (13732 bp)
chr scaffold424674 (10270 bp)
chr scaffold425050 (12114 bp)
chr scaffold425449 (12126 bp)
chr scaffold425469 (12533 bp)
chr scaffold428074 (12128 bp)
chr scaffold428188 (10736 bp)
chr scaffold431671 (12829 bp)
chr scaffold431819 (17424 bp)
chr scaffold431928 (10156 bp)
chr scaffold431960 (11632 bp)
chr scaffold432926 (11069 bp)
chr scaffold433332 (10628 bp)
chr scaffold434377 (13720 bp)
chr scaffold435031 (10705 bp)
chr scaffold436089 (10932 bp)
chr scaffold436358 (14110 bp)
chr scaffold436802 (11362 bp)
Number of paired-end alignments with a unique best hit: 490630
Mapping efficiency: 10.7%
Sequence pairs with no alignments under any condition: 4020912
Sequence pairs did not map uniquely: 90900
Sequence pairs which were discarded because genomic sequence could not be extracted: 89
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 247141 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 243400 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 490622
Mapping efficiency: 10.7%
Sequence pairs with no alignments under any condition: 4021272
Sequence pairs did not map uniquely: 90547
Sequence pairs which were discarded because genomic sequence could not be extracted: 84
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 247036 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 243502 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
Number of paired-end alignments with a unique best hit: 491111
Mapping efficiency: 10.7%
Sequence pairs with no alignments under any condition: 4020794
Sequence pairs did not map uniquely: 90537
Sequence pairs which were discarded because genomic sequence could not be extracted: 95
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 247821 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 243195 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.1 EPI-44_S41_L005_R2_001_val_2.fq.gz.temp.1 EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.2 EPI-44_S41_L005_R2_001_val_2.fq.gz.temp.2 EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.3 EPI-44_S41_L005_R2_001_val_2.fq.gz.temp.3
/home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005/EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005/EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_pe.bam /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005/EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_pe.bam
Number of paired-end alignments with a unique best hit: 1472363
Mapping efficiency: 10.7%
Sequence pairs with no alignments under any condition: 12062978
Sequence pairs did not map uniquely: 271984
Sequence pairs which were discarded because genomic sequence could not be extracted: 268
Number of sequence pairs with unique best (first) alignment came from the bowtie output:
CT/GA/CT: 741998 ((converted) top strand)
GA/CT/CT: 0 (complementary to (converted) top strand)
GA/CT/GA: 0 (complementary to (converted) bottom strand)
CT/GA/GA: 730097 ((converted) bottom strand)
Number of alignments to (merely theoretical) complementary strands being rejected in total: 0
EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.1_bismark_bt2_PE_report.txt EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.2_bismark_bt2_PE_report.txt EPI-44_S41_L005_R1_001_val_1.fq.gz.temp.3_bismark_bt2_PE_report.txt
################### Running MethylExtract v1.9 ###################
Functions summary:
Multi-fasta reference selected in ~/Documents/Genewiz_hdd/geo_genome/Panopea_generosa-Scaff-10k.fa
Watson strand FLAG: 99,147 & Crick strand FLAG: 83,163
Duplicated reads will not be deleted (default)
5' bases of the reads will not be ignored (default)
3' bases of the reads will not be ignored (default)
Pair-end overlapping parts will be included (default)
Quality values resume:
Selected Qscore: phred33-quals
Bisulfite check fraction/integer: 0.9
Bases from the 5' end to be ignored: 0
Bases from the 3' end to be ignored: 0
Minimum PHRED score for methylation and base call: 30
Minimum depth for methylation calls: 1
Minimum depth for SNV calls: 1
Alleles minimum frequency to be considered: 0.05
Maximum allowed strand bias: 0.7
SNV p-value threshold: 0.01
Activated Outputs:
Default methylation levels output: CG context
BED output format
WIG output format
SNVs VCF output format
Sorting alignments by chromosome
Reading /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005//EPI-44_S41_L005_R1_001_val_1_bismark_bt2_pe.bam
Starting main process
Merging Methylation & SNVs Outputs
Printing LOG file
# Methylation Ratios & SNVs retrieved, consult your results in /home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005/ #
# See the log file for further information #
zip error: Nothing to do! (EPI-44_S41_L005-sam/home/sean/Documents/Genewiz_hdd/EPI-44_S41_L005scaffold*.sam)
[1] "/home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005"
[1] "EPI-135WG_S42_L005_R1_001.fastq.gz"
[1] "EPI-135WG_S42_L005_R2_001.fastq.gz"Started analysis of EPI-135WG_S42_L005_R1_001.fastq.gz
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Approx 95% complete for EPI-135WG_S42_L005_R2_001.fastq.gzAnalysis complete for EPI-135WG_S42_L005_R2_001.fastq.gzNo quality encoding type selected. Assuming that the data provided uses Sanger encoded Phred scores (default)
Path to Cutadapt set as: 'cutadapt' (default)1.12Cutadapt seems to be working fine (tested command 'cutadapt --version')
AUTO-DETECTING ADAPTER TYPE
===========================
Attempting to auto-detect adapter type from the first 1 million sequences of the first file (>> /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R1_001.fastq.gz <<)
Found perfect matches for the following adapter sequences:
Adapter type Count Sequence Sequences analysed Percentage
Illumina 191466 AGATCGGAAGAGC 1000000 19.15
Nextera 0 CTGTCTCTTATA 1000000 0.00
smallRNA 0 TGGAATTCTCGG 1000000 0.00
Using Illumina adapter for trimming (count: 191466). Second best hit was Nextera (count: 0)
gzip: stdout: Broken pipe
Writing report to 'EPI-135WG_S42_L005_R1_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R1_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-135WG_S42_L005_R1_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R1_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
30000000 sequences processed
40000000 sequences processed
50000000 sequences processed
60000000 sequences processed
70000000 sequences processed
80000000 sequences processed
90000000 sequences processed
100000000 sequences processed
110000000 sequences processed
120000000 sequences processed
130000000 sequences processed
140000000 sequences processed
150000000 sequences processed
160000000 sequences processed
170000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R1_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 4035.94 s (23 us/read; 2.63 M reads/minute).
=== Summary ===
Total reads processed: 176,822,437
Reads with adapters: 92,962,555 (52.6%)
Reads written (passing filters): 176,822,437 (100.0%)
Total basepairs processed: 17,859,066,137 bp
Quality-trimmed: 63,031,675 bp (0.4%)
Total written (filtered): 16,384,399,522 bp (91.7%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 92962555 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 24.8%
C: 7.2%
G: 25.8%
T: 42.2%
none/other: 0.0%
Overview of removed sequences
length count expect max.err error counts
1 36226665 44205609.2 0 36226665
2 8653986 11051402.3 0 8653986
3 3115484 2762850.6 0 3115484
4 2117432 690712.6 0 2117432
5 1010080 172678.2 0 1010080
6 948159 43169.5 0 948159
7 866684 10792.4 0 866684
8 922361 2698.1 0 922361
9 942197 674.5 0 933381 8816
10 881709 168.6 1 842310 39399
11 923418 42.2 1 877239 46179
12 866404 10.5 1 825246 41158
13 863315 2.6 1 821357 41958
14 957253 2.6 1 902217 55036
15 919922 2.6 1 870735 49187
16 976739 2.6 1 919554 57185
17 946915 2.6 1 894247 52668
18 844876 2.6 1 802214 42662
19 895699 2.6 1 841228 54471
20 868341 2.6 1 823193 45148
21 943296 2.6 1 884900 58396
22 869528 2.6 1 823065 46463
23 852487 2.6 1 804803 47684
24 824852 2.6 1 775042 49810
25 788442 2.6 1 744484 43958
26 837638 2.6 1 784119 53519
27 786164 2.6 1 744300 41864
28 766789 2.6 1 727259 39530
29 809791 2.6 1 766405 43386
30 768263 2.6 1 730828 37435
31 806646 2.6 1 760318 46328
32 770705 2.6 1 731503 39202
33 784607 2.6 1 745124 39483
34 722497 2.6 1 687931 34566
35 706644 2.6 1 670600 36044
36 717165 2.6 1 678119 39046
37 816633 2.6 1 771302 45331
38 677255 2.6 1 643198 34057
39 616254 2.6 1 586847 29407
40 667522 2.6 1 631960 35562
41 969104 2.6 1 927744 41360
42 489199 2.6 1 465599 23600
43 400245 2.6 1 380185 20060
44 540321 2.6 1 513892 26429
45 544964 2.6 1 517263 27701
46 498483 2.6 1 473631 24852
47 538179 2.6 1 509414 28765
48 473605 2.6 1 447299 26306
49 513336 2.6 1 484703 28633
50 453441 2.6 1 430000 23441
51 447626 2.6 1 423553 24073
52 424424 2.6 1 400322 24102
53 389073 2.6 1 369067 20006
54 374286 2.6 1 353549 20737
55 402941 2.6 1 381040 21901
56 370348 2.6 1 349769 20579
57 336883 2.6 1 317486 19397
58 333543 2.6 1 316804 16739
59 322560 2.6 1 305993 16567
60 282823 2.6 1 268848 13975
61 305116 2.6 1 290022 15094
62 309118 2.6 1 293676 15442
63 262668 2.6 1 249439 13229
64 252432 2.6 1 240888 11544
65 227503 2.6 1 217695 9808
66 212203 2.6 1 202647 9556
67 206643 2.6 1 197441 9202
68 193664 2.6 1 185375 8289
69 194691 2.6 1 186978 7713
70 176965 2.6 1 169727 7238
71 172941 2.6 1 165778 7163
72 166033 2.6 1 158674 7359
73 186552 2.6 1 177180 9372
74 296800 2.6 1 287048 9752
75 170898 2.6 1 165695 5203
76 69028 2.6 1 66415 2613
77 40846 2.6 1 39175 1671
78 28004 2.6 1 26898 1106
79 17898 2.6 1 17108 790
80 14050 2.6 1 13462 588
81 9666 2.6 1 9220 446
82 7195 2.6 1 6855 340
83 5605 2.6 1 5352 253
84 3757 2.6 1 3571 186
85 2437 2.6 1 2306 131
86 1605 2.6 1 1513 92
87 1325 2.6 1 1236 89
88 1154 2.6 1 1064 90
89 1058 2.6 1 964 94
90 1252 2.6 1 1152 100
91 1512 2.6 1 1370 142
92 2045 2.6 1 1840 205
93 2925 2.6 1 2593 332
94 6201 2.6 1 5518 683
95 9025 2.6 1 8049 976
96 6494 2.6 1 5837 657
97 3586 2.6 1 3127 459
98 1198 2.6 1 1003 195
99 882 2.6 1 740 142
100 1024 2.6 1 799 225
101 2355 2.6 1 1556 799
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R1_001.fastq.gz
=============================================
176822437 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Writing report to 'EPI-135WG_S42_L005_R2_001.fastq.gz_trimming_report.txt'
SUMMARISING RUN PARAMETERS
==========================
Input filename: /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R2_001.fastq.gz
Trimming mode: paired-end
Trim Galore version: 0.4.2
Cutadapt version: 1.12
Quality Phred score cutoff: 20
Quality encoding type selected: ASCII+33
Adapter sequence: 'AGATCGGAAGAGC' (Illumina TruSeq, Sanger iPCR; auto-detected)
Maximum trimming error rate: 0.1 (default)
Minimum required adapter overlap (stringency): 1 bp
Minimum required sequence length for both reads before a sequence pair gets removed: 20 bp
Running FastQC on the data once trimming has completed
Running FastQC with the following extra arguments: ' -o ~/Documents/Genewiz_hdd/trimmed_fastqc/ '
Output file(s) will be GZIP compressed
Writing final adapter and quality trimmed output to EPI-135WG_S42_L005_R2_001_trimmed.fq.gz
>>> Now performing quality (cutoff 20) and adapter trimming in a single pass for the adapter sequence: 'AGATCGGAAGAGC' from file /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R2_001.fastq.gz <<<
10000000 sequences processed
20000000 sequences processed
30000000 sequences processed
40000000 sequences processed
50000000 sequences processed
60000000 sequences processed
70000000 sequences processed
80000000 sequences processed
90000000 sequences processed
100000000 sequences processed
110000000 sequences processed
120000000 sequences processed
130000000 sequences processed
140000000 sequences processed
150000000 sequences processed
160000000 sequences processed
170000000 sequences processed
This is cutadapt 1.12 with Python 2.7.12
Command line parameters: -f fastq -e 0.1 -q 20 -O 1 -a AGATCGGAAGAGC /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R2_001.fastq.gz
Trimming 1 adapter with at most 10.0% errors in single-end mode ...
Finished in 4232.03 s (24 us/read; 2.51 M reads/minute).
=== Summary ===
Total reads processed: 176,822,437
Reads with adapters: 108,587,123 (61.4%)
Reads written (passing filters): 176,822,437 (100.0%)
Total basepairs processed: 17,859,066,137 bp
Quality-trimmed: 121,398,456 bp (0.7%)
Total written (filtered): 16,364,304,059 bp (91.6%)
=== Adapter 1 ===
Sequence: AGATCGGAAGAGC; Type: regular 3'; Length: 13; Trimmed: 108587123 times.
No. of allowed errors:
0-9 bp: 0; 10-13 bp: 1
Bases preceding removed adapters:
A: 39.6%
C: 22.7%
G: 6.6%
T: 31.1%
none/other: 0.0%
Overview of removed sequences
length count expect max.err error counts
1 62472544 44205609.2 0 62472544
2 1371633 11051402.3 0 1371633
3 1152485 2762850.6 0 1152485
4 974650 690712.6 0 974650
5 977534 172678.2 0 977534
6 973069 43169.5 0 973069
7 932099 10792.4 0 932099
8 943799 2698.1 0 943799
9 921165 674.5 0 917761 3404
10 938059 168.6 1 904981 33078
11 888052 42.2 1 847489 40563
12 925139 10.5 1 885040 40099
13 880449 2.6 1 843090 37359
14 1007590 2.6 1 963625 43965
15 911402 2.6 1 873078 38324
16 929933 2.6 1 892039 37894
17 980541 2.6 1 940990 39551
18 819831 2.6 1 786118 33713
19 886089 2.6 1 850234 35855
20 878549 2.6 1 841131 37418
21 899233 2.6 1 858040 41193
22 927652 2.6 1 887568 40084
23 867353 2.6 1 832028 35325
24 919962 2.6 1 882018 37944
25 767000 2.6 1 734836 32164
26 793794 2.6 1 755792 38002
27 808715 2.6 1 764811 43904
28 857654 2.6 1 822816 34838
29 803691 2.6 1 763381 40310
30 909790 2.6 1 874944 34846
31 745546 2.6 1 711604 33942
32 806738 2.6 1 777804 28934
33 848967 2.6 1 812252 36715
34 824971 2.6 1 784638 40333
35 789215 2.6 1 763228 25987
36 726044 2.6 1 693877 32167
37 735579 2.6 1 706074 29505
38 629763 2.6 1 604392 25371
39 657120 2.6 1 629291 27829
40 648315 2.6 1 621519 26796
41 656135 2.6 1 633397 22738
42 641016 2.6 1 621445 19571
43 542551 2.6 1 520487 22064
44 564975 2.6 1 544103 20872
45 744311 2.6 1 722266 22045
46 507624 2.6 1 488372 19252
47 355254 2.6 1 339829 15425
48 525385 2.6 1 508747 16638
49 357928 2.6 1 344046 13882
50 387011 2.6 1 371612 15399
51 561705 2.6 1 545703 16002
52 306852 2.6 1 294546 12306
53 316942 2.6 1 303959 12983
54 276718 2.6 1 265122 11596
55 351562 2.6 1 339108 12454
56 334364 2.6 1 321079 13285
57 306889 2.6 1 295794 11095
58 285866 2.6 1 275563 10303
59 275468 2.6 1 264554 10914
60 268201 2.6 1 257762 10439
61 268357 2.6 1 257719 10638
62 277124 2.6 1 266243 10881
63 265820 2.6 1 255223 10597
64 258901 2.6 1 248418 10483
65 260494 2.6 1 250289 10205
66 260667 2.6 1 250646 10021
67 297320 2.6 1 284256 13064
68 653408 2.6 1 638481 14927
69 225009 2.6 1 216831 8178
70 104154 2.6 1 98845 5309
71 67588 2.6 1 63379 4209
72 56852 2.6 1 53131 3721
73 48657 2.6 1 45424 3233
74 40972 2.6 1 38306 2666
75 34813 2.6 1 32479 2334
76 30139 2.6 1 28060 2079
77 26533 2.6 1 24715 1818
78 22203 2.6 1 20618 1585
79 17429 2.6 1 16066 1363
80 13388 2.6 1 12378 1010
81 10044 2.6 1 9309 735
82 7571 2.6 1 7046 525
83 5410 2.6 1 4962 448
84 3341 2.6 1 3038 303
85 2048 2.6 1 1860 188
86 1253 2.6 1 1124 129
87 887 2.6 1 778 109
88 554 2.6 1 458 96
89 495 2.6 1 407 88
90 578 2.6 1 451 127
91 755 2.6 1 571 184
92 1304 2.6 1 1039 265
93 1984 2.6 1 1473 511
94 4321 2.6 1 3209 1112
95 6517 2.6 1 5011 1506
96 5115 2.6 1 4233 882
97 2754 2.6 1 2255 499
98 914 2.6 1 710 204
99 623 2.6 1 459 164
100 671 2.6 1 479 192
101 1685 2.6 1 1300 385
RUN STATISTICS FOR INPUT FILE: /home/sean/Documents/Genewiz_hdd/EPI-135WG_S42_L005_R2_001.fastq.gz
=============================================
176822437 sequences processed in total
The length threshold of paired-end sequences gets evaluated later on (in the validation step)
Validate paired-end files EPI-135WG_S42_L005_R1_001_trimmed.fq.gz and EPI-135WG_S42_L005_R2_001_trimmed.fq.gz
file_1: EPI-135WG_S42_L005_R1_001_trimmed.fq.gz, file_2: EPI-135WG_S42_L005_R2_001_trimmed.fq.gz
>>>>> Now validing the length of the 2 paired-end infiles: EPI-135WG_S42_L005_R1_001_trimmed.fq.gz and EPI-135WG_S42_L005_R2_001_trimmed.fq.gz <<<<<
Writing validated paired-end read 1 reads to EPI-135WG_S42_L005_R1_001_val_1.fq.gz
Writing validated paired-end read 2 reads to EPI-135WG_S42_L005_R2_001_val_2.fq.gz
Total number of sequences analysed: 176822437
Number of sequence pairs removed because at least one read was shorter than the length cutoff (20 bp): 489077 (0.28%)
>>> Now running FastQC on the validated data EPI-135WG_S42_L005_R1_001_val_1.fq.gz<<<
Started analysis of EPI-135WG_S42_L005_R1_001_val_1.fq.gz
Approx 5% complete for EPI-135WG_S42_L005_R1_001_val_1.fq.gz
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Approx 95% complete for EPI-135WG_S42_L005_R1_001_val_1.fq.gzAnalysis complete for EPI-135WG_S42_L005_R1_001_val_1.fq.gz
>>> Now running FastQC on the validated data EPI-135WG_S42_L005_R2_001_val_2.fq.gz<<<
Started analysis of EPI-135WG_S42_L005_R2_001_val_2.fq.gz
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Approx 60% complete for EPI-135WG_S42_L005_R2_001_val_2.fq.gzAdd a new chunk by clicking the Insert Chunk button on the toolbar or by pressing Ctrl+Alt+I.
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