RQ_dCt {RQdeltaCT}

RQ_dCt

Description

This function performs relative quantification of gene expression using 2^-dCt method.

Usage

RQ_dCt(
  data,
  group.study,
  group.ref,
  do.tests = TRUE,
  pairwise = FALSE,
  alternative = "two.sided",
  p.adjust.method = "BH",
  save.to.txt = FALSE,
  name.txt = "results_dCt"
)

Arguments

data

Data object returned from delta_Ct() or filter_transformed_data() function.

group.study

Character: name of study group (group of interest).

group.ref

Character: name of reference group.

do.tests

Logical: if TRUE, statistical significance of differences between compared groups will be calculated using Student's t test and Mann-Whitney U test. Default to TRUE.

pairwise

Logical: if TRUE, a pairwise analysis will be performed (see details). Default to FALSE.

alternative

Character: alternative hypothesis, must be one of "two.sided" (default), "greater" or "less".

p.adjust.method

Character: p value correction method for multiple testing, one of the "holm", "hochberg", "hommel", "bonferroni", "BH" (default), "BY","fdr", or "none". See documentation for stats::p.adjust() function for details.

save.to.txt

Logical: if TRUE, returned table with results will be saved to .txt file. Default to FALSE.

name.txt

Character: name of saved .txt file, without ".txt" name of extension. Default to "results_dCt".

Details

This function calculates: 1. Means (returned in columns with the "_mean" pattern) and standard deviations (returned in columns with the "_sd" pattern) of 2^-dCt transformed dCt values for each analyzed gene across compared groups. 2. P values of normality test (Shapiro_Wilk test) performed on 2^-dCt values across compared groups (returned in columns with the "_norm_p" pattern). 3. Fold change values (returned in "FCh" column) calculated for each gene by dividing mean of 2^-dCt values in study group by mean of 2^-dCt values in reference group. 4. Statistics (returned in column with the "_test_stat" pattern) and p values (returned in column with "_test_p" pattern) of differences in 2^-dCt values between study group and reference group using Student's t test and Mann-Whitney U test. 5. P values adjusted for multiple testing using a selected method.

Value

Data frame with results (if pairwise = FALSE) or, if pairwise = TRUE, the list object with two elements: a table with the results and the second table with fold change values calculated individually for each sample.

Examples

library(tidyverse)
library(coin)
data(data.Ct)
data.CtF <- filter_Ct(data.Ct,
                     remove.Gene = c("Gene2","Gene5","Gene6","Gene9","Gene11"),
                     remove.Sample = c("Control08","Control16","Control22"))
data.CtF.ready <- make_Ct_ready(data.CtF, imput.by.mean.within.groups = TRUE)
data.dCt <- delta_Ct(data.CtF.ready, ref = "Gene8", transform = TRUE)
results.dCt <- RQ_dCt(data.dCt,
                           group.study = "Disease",
                           group.ref = "Control")
head(results.dCt)