How to count RNA sequencereads using custom made python scripts? I am trying to do RNA seq analysis and my goal is to filter gene counts less than 5 using custom made python scripts. The code chunk goes as follows
#!/usr/bin/python
#Load relevant modules
import sys
    
#Read in the HTseq gene count output files for each of the 8 samples
inputlist = [sys.argv[1], sys.argv[2], sys.argv[3], sys.argv[4], sys.argv[5], sys.argv[6], sys.argv[7], sys.argv[8]]  
##Determine the genes with sufficient coverage
#Set up a variable to hold genes with sufficient coverage in any of the 8 samples
genelist = []

#For each HTseq file
for i in range(len(inputlist)):
    #Open it
    inputfile = open(inputlist[i], 'r')
    #For each line
    for line in inputfile:
        #Determine the number of reads/transcripts for this gene
        splitline = line.strip().split('\t')
        #Does it pass the threshold?
        if int(splitline[1]) &gt;= 5: 
            #If so, add it to list
            if not splitline[0] in genelist:
                genelist.append(splitline[0])
                    
    #Close the file
    inputfile.close()
    
#Write out the list of sufficiently expressed genes
outputlist = open(sys.argv[9], 'w') 
    
for gene in genelist:
    outputlist.write(gene + '\n')
        
outputlist.close()
##Filter each of the HTseq files for only sufficiently expressed genes across our samples
#For each HTseq file
    
for i in range(len(inputlist)):
    #Open it
    inputfile = open(inputlist[i], 'r')
    
#Open an output 'filtered' file
outputfile = open(sys.argv[i+10], 'w')
    
#For each line
for line in inputfile:
    #Determine the gene
    splitline = line.strip().split('\t')
        
    #Is it in our list?
    if splitline[0] in genelist:
        #If so, write out the original line
        outputfile.write(line)
            
#Close the files
inputfile.close()
outputfile.close()

python filtercounts.py 'wth1count.txt' 'wth2count.txt' 'wth3count.txt' 'wth4count.txt' '99h1count.txt' '99h2count.txt' '99h3count.txt' '99h4count.txt' 'genelist.txt' 'wth1filtered.txt' 'wth2filtered.txt' 'wth3filtered.txt' 'wth4filtered.txt'  '99h1filtered.txt' '99h2filtered.txt' '99h3filtered.txt' '99h4filtered.txt'
The program works fine but at the end of the code execution I am not getting the last two output files '99h3filtered.txt' '99h4filtered.txt'.
What software are you using for the next step?  DESeq in R will do this filtering for you in one line.
I think a more typical way to do this would be to make your entire gene count matrix, and then remove rows where the max value for that row is &lt; 5.  Your way, you have to go through each file twice; once to know which genes to keep, and then again to actually remove lines.