Curvas amplificación relativa final RT-qPCR
Aitana Barrientos
2026-04-28
Instalación de paquetes #Pacman: llama a otros paquetes y si no estan los instala
if(!require(pacman))
install.packages("pacman")## Loading required package: pacmanLlamar a pacman
library("pacman")Llamar paqueteria necesaria
p_load("vroom",
"dplyr",
"ggplot2",
"tidyr")Llamar a base de datos
Amplificacion_curves<- read.csv(file="https://raw.githubusercontent.com/ManuelLaraMVZ/Metabolomica_2026_1/refs/heads/main/Amplificaci%C3%B3n-RT-qPCR-20261_G1.csv")
Amplificacion_curves## Cycle A1 A2 A3 A4 A5
## 1 1 -9.4820849 -4.6293737 -11.7772746 -26.28535430 -5.8537227
## 2 2 -8.2355249 1.4191769 -8.0495902 -9.47855619 5.3672807
## 3 3 -1.6772250 2.0911121 -1.7854214 -5.30247480 3.9678107
## 4 4 0.3185735 5.6280553 7.2247356 1.14693074 8.5505417
## 5 5 2.2108334 0.8847123 2.8287136 -3.09043383 5.3771166
## 6 6 -6.6374424 -3.1625487 1.5613583 -6.03955623 5.9106199
## 7 7 -5.9423238 -0.7823888 -0.6270633 -1.03545902 0.8710510
## 8 8 1.9437281 3.6886636 0.2085411 0.62877114 1.3907730
## 9 9 -3.6448056 -1.1077201 -0.8618762 -2.95044851 -3.9775633
## 10 10 -0.9504983 -1.3818447 0.1964022 -3.44174230 -1.3140243
## 11 11 -1.0704692 -4.6268267 -6.0786860 -2.39480464 -1.9381592
## 12 12 -4.0826359 3.0639808 -5.1912493 -0.20388814 -5.3215744
## 13 13 -5.6351169 -2.5734721 4.3455890 1.38266219 -1.4750337
## 14 14 -0.1675053 8.6797492 9.0193333 4.20391947 3.7766950
## 15 15 -0.8401372 8.8392659 13.9957484 0.11106172 0.8799676
## 16 16 3.1419238 20.6178322 35.9797793 5.06223421 18.4882776
## 17 17 0.4453537 32.9489408 61.4085921 4.38789850 23.2535596
## 18 18 -0.6993381 58.6831283 90.5880275 0.09858559 45.7956588
## 19 19 -2.2119365 88.4611803 121.4411764 0.44823393 74.1387863
## 20 20 -1.0441537 119.0936698 148.4208196 7.17445771 106.9902667
## 21 21 0.3081392 144.9460662 170.4055821 1.60909720 135.6707376
## 22 22 2.3711248 163.0303977 189.9850745 2.42055684 157.9320331
## 23 23 0.1614810 175.6811662 207.8820747 -0.90989223 176.3156408
## 24 24 4.0503749 187.6572857 224.5392444 -1.36143150 191.2729995
## 25 25 4.7454935 199.6070647 238.3876641 2.53758775 204.6136745
## 26 26 7.8592963 210.2382783 250.9962958 2.03434321 214.2038484
## 27 27 3.1660928 217.1758991 261.4314367 2.14839117 221.5902889
## 28 28 5.9511283 219.3416774 268.5150038 9.08338713 225.8042541
## 29 29 11.8121388 222.4495320 273.7231170 0.40437049 229.7684731
## 30 30 4.6344001 225.7678660 278.7910732 0.88596480 232.4662385
## 31 31 4.4139032 231.9343416 285.4757579 1.49950508 236.0449098
## 32 32 3.2250317 235.3664922 290.2673024 0.06203012 235.8376688
## 33 33 3.6178860 241.6114577 292.5044770 0.67544777 236.1829262
## 34 34 3.3097533 243.5933785 294.1721172 2.70844373 234.4904944
## 35 35 5.8278075 247.0345531 295.3230749 -1.68708488 234.4634902
## 36 36 7.3028951 245.9013570 300.0756779 4.19819676 233.2749875
## 37 37 7.0780503 246.2882630 301.3884990 3.89055419 230.0798167
## 38 38 5.8847496 245.5842740 303.9718747 -1.67370839 223.9226030
## 39 39 3.9422104 246.9222362 304.0008804 -2.30902163 217.9624546
## 40 40 2.7576477 247.4540958 306.2909797 3.78478794 212.1123489
## 41 41 2.2611064 246.0142452 305.7459886 2.93997279 207.8086043
## 42 42 0.5248584 242.8084487 304.6009401 1.98881813 201.0294299
## 43 43 -1.7462855 239.9006451 303.1095185 0.49853976 196.6118218
## 44 44 -5.0249532 238.1537776 301.0707298 -4.79012379 189.1227225
## 45 45 -5.8501414 237.3050045 299.3908700 -3.70282106 184.6878919
## 46 46 -5.3429828 236.9909627 298.1232248 -3.03757552 180.1167596
## 47 47 -4.9894767 235.0602373 299.1819097 0.11330159 177.2809241
## 48 48 -6.4760990 232.7485888 297.1124623 -4.54477637 170.9702207
## 49 49 -5.6407421 230.4978762 296.1425288 -5.15423716 164.5240412
## 50 50 -5.4014170 230.5301180 294.9141860 -0.61060970 160.4963362
## 51 51 -4.5050870 230.5624039 296.8226743 1.35751066 159.3647416
## A6
## 1 -10.59438327
## 2 -0.92868366
## 3 -0.06897497
## 4 2.61675858
## 5 1.82031017
## 6 0.97070016
## 7 -1.82505540
## 8 3.64788428
## 9 0.45872975
## 10 2.31742306
## 11 -0.05761754
## 12 4.77083813
## 13 11.66457238
## 14 33.08126763
## 15 62.16627552
## 16 97.78390779
## 17 142.07676560
## 18 181.69907760
## 19 217.53589160
## 20 246.54191470
## 21 270.56598580
## 22 289.70480430
## 23 308.17328780
## 24 322.42669100
## 25 335.58013790
## 26 346.68773130
## 27 356.60789240
## 28 363.15883790
## 29 366.81331340
## 30 371.41880790
## 31 379.16585080
## 32 383.85857760
## 33 386.92169450
## 34 385.30511300
## 35 387.40891850
## 36 387.44410710
## 37 388.88151640
## 38 388.52667290
## 39 389.86346930
## 40 390.65261170
## 41 388.66381580
## 42 384.97947780
## 43 384.08587520
## 44 384.04249680
## 45 384.94550350
## 46 385.15613220
## 47 383.51805410
## 48 381.60343510
## 49 379.32131760
## 50 379.15061290
## 51 379.74683600Modificar base de datos #select: para seleccionar columnas #filter (una vez que ya conoces donde empieza la amplificación): para seleccionar filas
Amplificacion_curves2 <- Amplificacion_curves %>%
mutate(Ciclo = Cycle,
KAM = A1,
Muestra2 = A2,
Muestra3 = A3,
Muestra4 = A4,
Muestra5 = A5,
Muestra6 = A6) %>%
select(-Cycle:-A6)
Amplificacion_curves2## Ciclo KAM Muestra2 Muestra3 Muestra4 Muestra5
## 1 1 -9.4820849 -4.6293737 -11.7772746 -26.28535430 -5.8537227
## 2 2 -8.2355249 1.4191769 -8.0495902 -9.47855619 5.3672807
## 3 3 -1.6772250 2.0911121 -1.7854214 -5.30247480 3.9678107
## 4 4 0.3185735 5.6280553 7.2247356 1.14693074 8.5505417
## 5 5 2.2108334 0.8847123 2.8287136 -3.09043383 5.3771166
## 6 6 -6.6374424 -3.1625487 1.5613583 -6.03955623 5.9106199
## 7 7 -5.9423238 -0.7823888 -0.6270633 -1.03545902 0.8710510
## 8 8 1.9437281 3.6886636 0.2085411 0.62877114 1.3907730
## 9 9 -3.6448056 -1.1077201 -0.8618762 -2.95044851 -3.9775633
## 10 10 -0.9504983 -1.3818447 0.1964022 -3.44174230 -1.3140243
## 11 11 -1.0704692 -4.6268267 -6.0786860 -2.39480464 -1.9381592
## 12 12 -4.0826359 3.0639808 -5.1912493 -0.20388814 -5.3215744
## 13 13 -5.6351169 -2.5734721 4.3455890 1.38266219 -1.4750337
## 14 14 -0.1675053 8.6797492 9.0193333 4.20391947 3.7766950
## 15 15 -0.8401372 8.8392659 13.9957484 0.11106172 0.8799676
## 16 16 3.1419238 20.6178322 35.9797793 5.06223421 18.4882776
## 17 17 0.4453537 32.9489408 61.4085921 4.38789850 23.2535596
## 18 18 -0.6993381 58.6831283 90.5880275 0.09858559 45.7956588
## 19 19 -2.2119365 88.4611803 121.4411764 0.44823393 74.1387863
## 20 20 -1.0441537 119.0936698 148.4208196 7.17445771 106.9902667
## 21 21 0.3081392 144.9460662 170.4055821 1.60909720 135.6707376
## 22 22 2.3711248 163.0303977 189.9850745 2.42055684 157.9320331
## 23 23 0.1614810 175.6811662 207.8820747 -0.90989223 176.3156408
## 24 24 4.0503749 187.6572857 224.5392444 -1.36143150 191.2729995
## 25 25 4.7454935 199.6070647 238.3876641 2.53758775 204.6136745
## 26 26 7.8592963 210.2382783 250.9962958 2.03434321 214.2038484
## 27 27 3.1660928 217.1758991 261.4314367 2.14839117 221.5902889
## 28 28 5.9511283 219.3416774 268.5150038 9.08338713 225.8042541
## 29 29 11.8121388 222.4495320 273.7231170 0.40437049 229.7684731
## 30 30 4.6344001 225.7678660 278.7910732 0.88596480 232.4662385
## 31 31 4.4139032 231.9343416 285.4757579 1.49950508 236.0449098
## 32 32 3.2250317 235.3664922 290.2673024 0.06203012 235.8376688
## 33 33 3.6178860 241.6114577 292.5044770 0.67544777 236.1829262
## 34 34 3.3097533 243.5933785 294.1721172 2.70844373 234.4904944
## 35 35 5.8278075 247.0345531 295.3230749 -1.68708488 234.4634902
## 36 36 7.3028951 245.9013570 300.0756779 4.19819676 233.2749875
## 37 37 7.0780503 246.2882630 301.3884990 3.89055419 230.0798167
## 38 38 5.8847496 245.5842740 303.9718747 -1.67370839 223.9226030
## 39 39 3.9422104 246.9222362 304.0008804 -2.30902163 217.9624546
## 40 40 2.7576477 247.4540958 306.2909797 3.78478794 212.1123489
## 41 41 2.2611064 246.0142452 305.7459886 2.93997279 207.8086043
## 42 42 0.5248584 242.8084487 304.6009401 1.98881813 201.0294299
## 43 43 -1.7462855 239.9006451 303.1095185 0.49853976 196.6118218
## 44 44 -5.0249532 238.1537776 301.0707298 -4.79012379 189.1227225
## 45 45 -5.8501414 237.3050045 299.3908700 -3.70282106 184.6878919
## 46 46 -5.3429828 236.9909627 298.1232248 -3.03757552 180.1167596
## 47 47 -4.9894767 235.0602373 299.1819097 0.11330159 177.2809241
## 48 48 -6.4760990 232.7485888 297.1124623 -4.54477637 170.9702207
## 49 49 -5.6407421 230.4978762 296.1425288 -5.15423716 164.5240412
## 50 50 -5.4014170 230.5301180 294.9141860 -0.61060970 160.4963362
## 51 51 -4.5050870 230.5624039 296.8226743 1.35751066 159.3647416
## Muestra6
## 1 -10.59438327
## 2 -0.92868366
## 3 -0.06897497
## 4 2.61675858
## 5 1.82031017
## 6 0.97070016
## 7 -1.82505540
## 8 3.64788428
## 9 0.45872975
## 10 2.31742306
## 11 -0.05761754
## 12 4.77083813
## 13 11.66457238
## 14 33.08126763
## 15 62.16627552
## 16 97.78390779
## 17 142.07676560
## 18 181.69907760
## 19 217.53589160
## 20 246.54191470
## 21 270.56598580
## 22 289.70480430
## 23 308.17328780
## 24 322.42669100
## 25 335.58013790
## 26 346.68773130
## 27 356.60789240
## 28 363.15883790
## 29 366.81331340
## 30 371.41880790
## 31 379.16585080
## 32 383.85857760
## 33 386.92169450
## 34 385.30511300
## 35 387.40891850
## 36 387.44410710
## 37 388.88151640
## 38 388.52667290
## 39 389.86346930
## 40 390.65261170
## 41 388.66381580
## 42 384.97947780
## 43 384.08587520
## 44 384.04249680
## 45 384.94550350
## 46 385.15613220
## 47 383.51805410
## 48 381.60343510
## 49 379.32131760
## 50 379.15061290
## 51 379.74683600Reorganizar datos
Amplificacion_curves3 <- Amplificacion_curves2 %>%
pivot_longer(cols = -Ciclo,
names_to = "Muestras",
values_to = "Fluorescencia")
Amplificacion_curves3## # A tibble: 306 × 3
## Ciclo Muestras Fluorescencia
## <int> <chr> <dbl>
## 1 1 KAM -9.48
## 2 1 Muestra2 -4.63
## 3 1 Muestra3 -11.8
## 4 1 Muestra4 -26.3
## 5 1 Muestra5 -5.85
## 6 1 Muestra6 -10.6
## 7 2 KAM -8.24
## 8 2 Muestra2 1.42
## 9 2 Muestra3 -8.05
## 10 2 Muestra4 -9.48
## # ℹ 296 more rowsGráfica de amplificación
Grafica_amplificacion <- ggplot(Amplificacion_curves3,
aes(x = Ciclo,
y = Fluorescencia,
color = Muestras)) +
geom_line(linewidth = 1.5) +
theme_classic() +
labs(title = "Curvas de amplificación de valores relativos RT-qPCR",
subtitle = "Todas las muestras",
caption = "Equipo: KAM",
x = "Ciclo",
y = "Fluorescencia (u.a.)") +
theme(axis.line = element_line(size = 1.2, color = "black"),
axis.title = element_text(face = "bold"),
axis.text = element_text(face = "bold"),
legend.title = element_text(face = "bold"),
legend.text = element_text(face = "bold"))## Warning: The `size` argument of `element_line()` is deprecated as of ggplot2 3.4.0.
## ℹ Please use the `linewidth` argument instead.
## This warning is displayed once per session.
## Call `lifecycle::last_lifecycle_warnings()` to see where this warning was
## generated.Grafica_amplificacion
Definir umbral de ciclos y mejorar la gráfica
# Definir umbral de ciclos
Umbral_ciclos <- 0.008
# Filtrar los ciclos de interés (ejemplo: del 9 al 35)
Amplificacion_curves_filtrado <- Amplificacion_curves3 %>%
filter(Ciclo >= 9 & Ciclo <= 35)
# Gráfica mejorada con umbral y zoom
Grafica_amplif2 <- ggplot(Amplificacion_curves_filtrado,
aes(x = Ciclo,
y = Fluorescencia,
color = Muestras)) +
geom_line(linewidth = 1.5) +
geom_hline(yintercept = Umbral_ciclos, linetype = "dashed", color = "#581845") +
theme_classic() +
labs(title = "Curvas de amplificación de valores relativos RT-qPCR",
subtitle = "Ciclos 9–35",
caption = "Equipo: KAM",
x = "Ciclos",
y = "Fluorescencia (u.a.)") +
theme(axis.line = element_line(linewidth = 1.2, color = "black"),
axis.title = element_text(face = "bold"),
axis.text = element_text(face = "bold"),
legend.title = element_text(face = "bold"),
legend.text = element_text(face = "bold")) +
scale_x_continuous(breaks = seq(min(Amplificacion_curves_filtrado$Ciclo),
max(Amplificacion_curves_filtrado$Ciclo),
by = 2)) +
scale_y_continuous(labels = scales::number_format(accuracy = 0.02))
Grafica_amplif2
Gráfica Equipo KAM
Grafica_amplif_equipo <- ggplot(Amplificacion_curves %>% filter(Cycle >= 9 & Cycle <= 35),
aes(x = Cycle)) +
geom_line(aes(y = A1, color = "KAM"), linewidth = 1.5) +
geom_line(aes(y = A6, color = "A3"), linewidth = 1.5) +
geom_hline(yintercept = Umbral_ciclos, linetype = "dashed", color = "#581845") +
scale_color_manual(values = c("KAM" = "#1e2f7c", "A3" = "#900C3F")) +
theme_classic() +
labs(title = "Curvas de amplificación de valores relativos RT-qPCR",
subtitle = "Muestra KAM vs Muestra Equipo A3",
caption = "Equipo: KAM",
x = "Ciclos",
y = "Fluorescencia (u.a.)",
color = "Muestras") +
theme(axis.line = element_line(linewidth = 1.2, color = "black"),
axis.title = element_text(face = "bold"),
axis.text = element_text(face = "bold"),
legend.title = element_text(face = "bold"),
legend.text = element_text(face = "bold")) +
scale_x_continuous(breaks = seq(1, 35, by = 2))
Grafica_amplif_equipo