QPCR结果R语言绘图

library(tidyverse)
qpcr_count <- function(raw_data,ref_sample,ref_gene) {
  colnames(raw_data) = c("Sample Name","Target Name","CT" )
  raw_data$CT = raw_data$CT %>% as.numeric()
  
  Treat_samples = raw_data$`Sample Name` %>% unique() %>% setdiff(ref_sample)
  genes = raw_data$`Target Name` %>% unique() %>%  setdiff(ref_gene)
  
  
  result = raw_data %>%left_join(
    raw_data %>%  
      filter(`Target Name` == ref_gene) %>% 
      group_by(`Sample Name`) %>% 
      mutate(mean_ref_gene = mean(CT)) %>% 
      dplyr::select(`Sample Name`,mean_ref_gene) %>% 
      unique(),
    by =c("Sample Name")
  ) %>% 
    mutate(DeltaCT = CT - mean_ref_gene)  %>% 
    left_join(
      raw_data %>%left_join(
        raw_data %>%  
          filter(`Target Name` == ref_gene) %>% 
          group_by(`Sample Name`) %>% 
          mutate(mean_ref_gene = mean(CT)) %>% 
          dplyr::select(`Sample Name`,mean_ref_gene) %>% 
          unique(),
        by =c("Sample Name")
      ) %>% 
        mutate(DeltaCT = CT - mean_ref_gene)  %>% 
        filter(`Sample Name` == ref_sample) %>% group_by(`Target Name`) %>% 
        mutate(CK_sample_mean_DeltaCT = mean(DeltaCT)) %>%  ##  分别计算对照组每个基因的DeltaCT的均值
        dplyr::select(`Target Name`,CK_sample_mean_DeltaCT) %>% 
        unique(),by =c("Target Name")
    ) %>%  
    mutate(DeltaDeltaCT = DeltaCT - CK_sample_mean_DeltaCT) %>%  ## 计算DeltaDeltaCT
    mutate(RelativeExpression = 2^(-DeltaDeltaCT))  %>% ## 计算2^(-DeltaDeltaCT)
    dplyr::select("Sample Name","Target Name","CT","DeltaCT" ,"DeltaDeltaCT","RelativeExpression" )
  
  return(result)
}

library(readxl)
qocr <- read_excel("~/Program/wanggang/螺/qocr.xlsx")

qocr$`Sample Name` = qocr$`Sample Name` %>% str_sub(1,1)

qpcr_data =qocr

qpcr_res = qpcr_data %>%qpcr_count(ref_sample = "L",ref_gene = "ACTIN")


qpcr_res_sum =qpcr_data %>% 
  qpcr_count(ref_sample = "L",ref_gene = "ACTIN") %>% 
  filter(`Sample Name` == "L") %>% 
  group_by(`Target Name`) %>% 
  summarise(mean = mean(RelativeExpression),
            sd = sd(RelativeExpression)) %>% 
  mutate(`Sample Name` = "L") %>% 
  bind_rows(
    qpcr_data %>% 
      qpcr_count(ref_sample = "L",ref_gene = "ACTIN")%>% 
      filter(`Sample Name` != "L") %>% 
      group_by(`Target Name`) %>% 
      summarise(mean = mean(RelativeExpression),
                sd = sd(RelativeExpression)) %>% 
      mutate(`Sample Name` = "J") 
  )


qpcr_res_sum$`Sample Name` = qpcr_res_sum$`Sample Name` %>% factor(levels = c("L","J"))
qpcr_res$`Sample Name`= qpcr_res$`Sample Name` %>% factor(levels = c("L","J"))

c("ACTIN","SOX2","SOX10","WNT2","WNT7","FOXL2")

qpcr_res_sum$`Sample Name` = qpcr_res_sum$`Sample Name` %>% str_replace_all("L","ovaries") %>% str_replace_all("J","testes")
qpcr_res$`Sample Name` = qpcr_res$`Sample Name` %>% str_replace_all("J","testes")%>% str_replace_all("L","ovaries") 

genenames = qpcr_res_sum$`Target Name` %>% unique() %>% setdiff("ACTIN")

qpcr_res_sum1 = qpcr_res_sum %>% filter(`Target Name` %in% c("ACTIN",genenames[1:5]) )
qpcr_res1 = qpcr_res %>%  filter(`Target Name` %in% c("ACTIN",genenames[1:5]) )

library(ggsignif)
p1 = ggplot(data = qpcr_res_sum1 ,
            aes(x = `Sample Name` , y = mean))+
  
  geom_col(
    aes(x = `Sample Name` , y = mean,fill = `Sample Name`),
    width = 0.7)+
  geom_errorbar(aes(ymin = mean - sd,
                    ymax = mean + sd),
                fill =  "black",
                width = 0.15)+
  geom_signif(data = qpcr_res1,
              aes(x = `Sample Name`,
                  y = RelativeExpression),
              comparisons = list(c("ovaries", "testes")),
              test = "t.test",
              map_signif_level = F   #是否显示*号代表差异性
  )+
  scale_fill_manual(values = c('royalblue',  '#FF0124')) +
  theme_bw()+
  facet_grid(. ~ `Target Name` ) +
  theme(legend.position = "none") +
  labs(x = NULL, y = "RelativeExpression") +
  theme(axis.title = element_text(size = 12, face = "bold"), axis.text = element_text(face = "bold",colour = "gray0"),
        plot.title = element_text(size = 15)) + theme(plot.title = element_text(face = "bold"),
                                                      legend.text = element_text(face = "bold"),
                                                      legend.title = element_text(face = "bold"),
                                                      legend.position = "top", legend.direction = "horizontal") +
  labs(fill = "type")