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Combining RNA-Seq read counts from 2 lanes of the same sample (.txt file)

Hi,

I have a question on combining read counts from 2 lanes of the same sample. I have a very large RNA-Seq dataset downloaded from the NCBI GEO. The data files are in the *.txt format and each sample with 2 lanes obtained from featureCounts. I would like to perform the differential expression analysis using this data. Since, each sample is of 2 lanes, please let me know if these samples can be combined by sum or mean or if there is any other method?

dput(Sample_1_L1_S65_L001_R1_001Aligned.sortedByCoord.out_gene.featureCounts.txt)

structure(list(Geneid = c("ENSG00000223972", "ENSG00000227232", 
                          "ENSG00000278267", "ENSG00000243485", "ENSG00000237613", "ENSG00000268020", 
                          "ENSG00000240361", "ENSG00000186092"), Sample_1_L1_S65_L001_R1_001Aligned.sortedByCoord.out.bam.txt = c(0L, 
                                                                                                                              22L, 1L, 0L, 0L, 4L, 0L, 6L)), class = "data.frame", row.names = c(NA, 
                                                                                                                                                                                                 -8L))
#>            Geneid Sample_1_L1_S65_L001_R1_001Aligned.sortedByCoord.out.bam.txt
#> 1 ENSG00000223972                                                            0
#> 2 ENSG00000227232                                                           22
#> 3 ENSG00000278267                                                            1
#> 4 ENSG00000243485                                                            0
#> 5 ENSG00000237613                                                            0
#> 6 ENSG00000268020                                                            4
#> 7 ENSG00000240361                                                            0
#> 8 ENSG00000186092                                                            6

dput(Sample_1_L1_S65_L002_R1_001Aligned.sortedByCoord.out_gene.featureCounts.txt)

structure(list(Geneid = c("ENSG00000223972", "ENSG00000227232", 
                          "ENSG00000278267", "ENSG00000243485", "ENSG00000237613", "ENSG00000268020", 
                          "ENSG00000240361", "ENSG00000186092"), Sample_1_L1_S65_L002_R1_001Aligned.sortedByCoord.out.bam.txt = c(1L, 
                                                                                                                              27L, 2L, 0L, 0L, 2L, 0L, 1L)), class = "data.frame", row.names = c(NA, 
                                                                                                                                                                                                 -8L))
#>            Geneid Sample_1_L1_S65_L002_R1_001Aligned.sortedByCoord.out.bam.txt
#> 1 ENSG00000223972                                                            1
#> 2 ENSG00000227232                                                           27
#> 3 ENSG00000278267                                                            2
#> 4 ENSG00000243485                                                            0
#> 5 ENSG00000237613                                                            0
#> 6 ENSG00000268020                                                            2
#> 7 ENSG00000240361                                                            0
#> 8 ENSG00000186092                                                            1

Created on 2021-09-18 by the [reprex package](https://reprex.tidyverse.org) (v2.0.1)

Thank you,

Toufiq

expression featurecounts counts rna-seq gene

1 answer

Simply add the counts from the technical replicates, for example by using the sumTechReps function in the edgeR package, which is provided for this purpose. Taking means is not appropriate.

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