Fastqc Summary

Pre-trimmed seqs

ID Index_# Library_prep_selection Total Sequences
M04 9 Pippin 44341403
M05 10 Pippin 27033086
3.204.P 12 Pippin 16330176
M11 11 Pippin 15606044
1.215 1 Gel (Montgomery Lab) 38591844
1.204B 2 Gel (Montgomery Lab) 35851019
1.201B.P 3 Gel (Montgomery Lab) 36650445
1.206B 4 Gel (Montgomery Lab) 34146111
1.218.P 5 Gel (Montgomery Lab) 33330352
2.208B 6 Gel (Montgomery Lab) 45455664
3.208B.P 7 Gel (Montgomery Lab) 21343664
4.211B.P 8 Gel (Montgomery Lab) 37541519
Undetermined _ _ 28100877
Totals _ _ 414322204

Adapter and Quality Trimming

bbduk.sh for adapter and quality trimming

bbduk.sh -Xmx1g in=$file out=Trim_$file ref=adapters,phix ktrim=r k=23 mink=11 hdist=2 qtrim=rl trimq=10 ftm=5 maq=15 tpe tbo

  # ref: specify reference files for illumina adapters and PhiX contaminants
  # k: kmer length
  # qtrim: right and left side of read qual trim
  # trimq: phred trim algorithm, phred algorithm [...] leaves the longest area with average quality 
  # above the limit you specify such that it cannot be extended without adding an area of average quality 
  # below the limit you specify
  # ftm: clips the last base of every read, 76bp -> 75bp
  # maq: filter, removes all reads with average quality below value
ID Index_# Library_prep_selection Organ Total_sequences Post_trim_total_sequences Percent_remaining
M04 9 Pippin Brain 44341403 37217334 0.839336
M05 10 Pippin Brain 27033086 19547797 0.7231064
3.204.P 12 Pippin Ovary 16330176 13665024 0.8367959
M11 11 Pippin Brain 15606044 8415387 0.539239
1.215 1 Gel (Montgomery Lab) Ovary 38591844 28899896 0.7488602
1.204B 2 Gel (Montgomery Lab) Ovary 35851019 27808096 0.7756571
1.201B.P 3 Gel (Montgomery Lab) Ovary 36650445 32605999 0.8896481
1.206B 4 Gel (Montgomery Lab) Ovary 34146111 28993524 0.8491018
1.218.P 5 Gel (Montgomery Lab) Ovary 33330352 29422636 0.882758
2.208B 6 Gel (Montgomery Lab) Ovary 45455664 37283314 0.8202127
3.208B.P 7 Gel (Montgomery Lab) Ovary 21343664 16204049 0.7591972
4.211B.P 8 Gel (Montgomery Lab) Ovary 37541519 29223604 0.7784342
Undetermined _ _ _ 28100877 10095199 0.3592485
Totals _ _ _ 414322204 282164525 0.6810268

Example sample: 1.201B (ovary)

———————————— Pre-trim ———————————————————————– Post-trim —————————-

Example sample: M05 (brain)

———————————— Pre-trim ———————————————————————– Post-trim —————————-

5p Read Uracil Enrichment

Sample: 1.201B (ovary)

Sample: M05 (brain)

Mapping - bbmap.sh

Examples: 1.201B and M05
bbmap.sh in=Trim_1_201B_P_S7_R1_001.fastq.gz out=1_201B.bam covstats=covstats.txt scafstats=scafstats.txt 1>>log.txt 2>&1

bbmap.sh in=Trim_M05_S2_R1_001.fastq.gz out=M05.bam.gz covstats=M05_covstats.txt scafstats=M05_scafstats.txt 1>>log.txt 2>&1

Mapping Results

Reads Used: 32605999 (717474644 bases)

1.201B pct reads num reads pct bases num bases
mapped: 72.7929% 23734866 70.7780% 507814367
unambiguous: 14.6013% 4760912 16.4424% 117969709
ambiguous: 58.1916% 18973954 54.3357% 389844658
perfect best site: 32.7053% 10663900 32.0934% 230262179
semiperfect site: 33.1714% 10815859 32.6536% 234281292

Reads Used: 19547797 (389132393 bases)

M05 pct reads num reads pct bases num bases
mapped: 84.5843% 16534370 87.9472% 342231195
unambiguous: 8.7994% 1720095 10.1004% 39303964
ambiguous: 75.7849% 14814275 77.8468% 302927231
perfect best site: 44.0712% 8614941 44.3583% 172612591
semiperfect site: 44.1723% 8634713 44.4622% 173016816