Curvas de disociación RT-qPCR

if(!require(pacman))
install.packages("pacman")

## Loading required package: pacman

## Loading required package: pacman
library("pacman")
install.packages("dplyr")

## Installing package into '/cloud/lib/x86_64-pc-linux-gnu-library/4.4'
## (as 'lib' is unspecified)

## Installing package into '/cloud/lib/x86_64-pc-linux-gnu-library/4.4'
## (as 'lib' is unspecified)
library("dplyr")

## 
## Attaching package: 'dplyr'

## The following objects are masked from 'package:stats':
## 
##     filter, lag

## The following objects are masked from 'package:base':
## 
##     intersect, setdiff, setequal, union

p_load("vroom",
       "dplyr",
       "ggplot2",
       "tidyr",
       "ggrepel",
       "plotly")

Melting_curves <- vroom("https://raw.githubusercontent.com/ManuelLaraMVZ/resultados_PCR_practica/refs/heads/main/Disociaci%C3%B3n_Grupo1_17022024.csv")

## Rows: 61 Columns: 7
## ── Column specification ────────────────────────────────────────────────────────
## Delimiter: ","
## dbl (7): Temperature, A1, B1, C1, D1, E1, F1
## 
## ℹ Use `spec()` to retrieve the full column specification for this data.
## ℹ Specify the column types or set `show_col_types = FALSE` to quiet this message.

Melting_curves

modificar la base de datos

Melting_curves2 <- Melting_curves %>% 
  mutate(Temperatura = Temperature,PPDA = A1, ZARX = B1, F2S2 = C1, LANS =D1, NEGATIVO = E1, D2MI = F1) %>% 
  select(-Temperature:-F1) 
  
Melting_curves2

Agrupacion de datos

Melting_curves3 <- Melting_curves2 %>% 
  pivot_longer(cols = -Temperatura,
               names_to = "Muestras",
               values_to = "Fluorescencias")
Melting_curves3

Grafica

Grafica_melting <- ggplot(Melting_curves3,
                          aes(x=Temperatura,
                              y=Fluorescencias,
                              color=Muestras))+
  geom_line(linewidth=1.5)+
    labs(title = "Curvas de disociación",
       subtitle = "Todas las muestras",
       caption = "Diseñó: zulia, renata, ximena, ana gaby",
       x="Temperatura(ºC)",
       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 every 8 hours.
## Call `lifecycle::last_lifecycle_warnings()` to see where this warning was
## generated.

Grafica_melting

Derivadas de las curvas de disociacion

Derivadas <- Melting_curves2 %>% 
  mutate(across(PPDA:D2MI,
                ~-c(NA, diff(.x)/diff(Melting_curves2$Temperatura)), .names = "d_{.col}")) %>% 
  select(-PPDA: -D2MI) %>% 
  slice(-1)
Derivadas

Reordenar los datos

Derivadas2 <- Derivadas %>% 
  pivot_longer(cols = -Temperatura,
               names_to = "Muestras",
               values_to = "Derivadas")
Derivadas2

Graficas Derivadas

Grafica_derivada <- ggplot(Derivadas2,
                           aes(x=Temperatura,
                               y=Derivadas,
                               color=Muestras))+
   geom_line(linewidth=1.5)+
  theme_classic( )+
  labs(title = "Curvas de la derivada de disociación",
       subtitle = "Todas las muestras",
       caption = "Diseñó: zulia, renata, ximena, ana gaby",
       x="Temperatura(ºC)",
       y=expression(-Delta-F/Delta-T))+
  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"))+
scale_x_continuous(breaks=seq(min(Derivadas2$Temperatura),
                       max(Derivadas2$Temperatura),
                       by=2.5))
Grafica_derivada

Construir una gráfica 3D

Grafica_derivada_3D <- plot_ly(Derivadas2,
                               x= ~Temperatura,
                               y= ~Muestras,
                               z= ~Derivadas,
                               color=~factor(Muestras),
                               type="scatter3d",
                               mode="lines",
                               line=list(width=6)) %>% 
  layout(title="Curva de derivada en 3D",
         scene=list(),
         xaxis=list(title="Temperarura(ºC)"),
         yaxis=list(title="Muestras"),
         zaxis=list(title="-ΔT/ΔF"))
Grafica_derivada_3D

## Warning: 'layout' objects don't have these attributes: 'zaxis'
## Valid attributes include:
## '_deprecated', 'activeshape', 'annotations', 'autosize', 'autotypenumbers', 'calendar', 'clickmode', 'coloraxis', 'colorscale', 'colorway', 'computed', 'datarevision', 'dragmode', 'editrevision', 'editType', 'font', 'geo', 'grid', 'height', 'hidesources', 'hoverdistance', 'hoverlabel', 'hovermode', 'images', 'legend', 'mapbox', 'margin', 'meta', 'metasrc', 'modebar', 'newshape', 'paper_bgcolor', 'plot_bgcolor', 'polar', 'scene', 'selectdirection', 'selectionrevision', 'separators', 'shapes', 'showlegend', 'sliders', 'smith', 'spikedistance', 'template', 'ternary', 'title', 'transition', 'uirevision', 'uniformtext', 'updatemenus', 'width', 'xaxis', 'yaxis', 'barmode', 'bargap', 'mapType'

Grafica del equipo

Grafica_derivada_equipo <- ggplot(Derivadas,
                                  aes(x=Temperatura))+
geom_line(aes(y= d_ZARX, color = "d_ZARX"), linewidth = 1.2) + # Leyenda para la línea ZARX
geom_line(aes(y= d_NEGATIVO, color="d_NEGATIVO"),linewidth=1.2)+
scale_color_manual(values = c("d_ZARX" = "#ce9df2", "d_NEGATIVO" = "#49c1e1"))+
theme_classic()+
labs(title = "Derivadas de disociación",
     subtitle = "Tejido:Cerebro",
     caption= "Diseñó: Zulia, Renata, Ximena, Ana Gaby",
     x="Temperatura",
 y=expression(-Delta-F/Delta-T))+
  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"))+
scale_x_continuous(breaks=seq(min(Derivadas2$Temperatura),
                       max(Derivadas2$Temperatura),
                       by=2.5))
Grafica_derivada_equipo

Etiquetas

Pico <- max(Derivadas$d_ZARX)
Pico

## [1] 43.78384

Temperatura

Tm <- Derivadas %>% 
    filter(d_ZARX==Pico) %>% 
  select(Temperatura, d_ZARX)
Tm

Grafica con etiqueta

Grafica_derivada_equipo2 <- Grafica_derivada_equipo+
  geom_vline(xintercept = Tm$Temperatura, color="#ce9df2", 
             linetype="dashed",
             linewidth=1)+
  geom_label_repel(data=Tm,
                   aes(x=Temperatura,
                       y= d_ZARX,
                       label=paste("Tm=", round(Temperatura),"ºC")),
                   nudge_x=4,
                   nudge_y = -0.03,
                   max.overlaps = 100,
                   color="black",
                   fill="#ce9df2")
Grafica_derivada_equipo2