how to count the nucleotide frequency of paired ended fasta files

Question

[Learner]

I have read this Counting nucleotide frequency using perl script which is very interesting. however, the fastq files are not in this format and I would like to know if there is any avaable code for paired ended sequencing

[Friederike]

You need to be a little more specific:

• What exactly do you want to achieve?
• What type of data do you have?
• Why do you think the script is not doing what you need, i.e. do you have an error message or example output that illustrates why you think it's failing you?

[Learner]

@Friederike

So cliche but I try to answer What exactly do you want to achieve? achieving to read the nucleotide frequency What type of data do you have? fastq paired ended Why do you think the script is not doing what you need, I think you need to know how the structure of a paired ended is !!!

sorry but your questions are not useful whatsoever

[Friederike]

well, the original script you linked to was for fastq files and paired-end sequencing is typically simply dumped in two separate fastq files, so I did not understand why that script should not be working. I admit I did not fully read the title of your post, only the content which started with "This script is interesting". We might be able to point you to better solutions if you told us, for example, if you needed a specific table output or a plot (such as this and so on.... Anyway, I shall refrain from further unhelpful comments :)

[Learner]

@Friederike comments is always good as long as one does not send another for chasing a white goose ! or just blah blah. I try to be very sharp in my questions so that I don't waste people time and sorry if I am not perfect. Yes there are two Fasta and not fastq ! it is just the matter of structure of the data , I am trying to figure out

[GenoMax]

the fastq files are not in this format

That is easily fixed using reformat.sh from BBMap suite.

reformat.sh in=R1.fq.gz out=R1.fa

On a serious note you could also look at this: https://digibio.blogspot.com/2017/12/nucleotide-base-frequency-per-read-and.html

[Learner]

@genomax can you paste the reformat.sh here please? or is it this bash file ? https://github.com/BioInfoTools/BBMap/blob/master/sh/reformat.sh

[GenoMax]

Official BBMap repo is located here. Reformat is a versatile utility that does a ton of other things. You can find a guide here. It is part of a much bigger BBTools package.

[Learner]

@genomax , do you know what exactly it does to each fastq ? I means I should do that for both forward and reverse right ?

Thanks

[GenoMax]

If you were planning to use the fasta perl script linked in your original post reformat command above will convert your fastq format reads into fasta format. You will have to do it for F/R reads.

[Damian Kao]

Are your fastqs paired in the same order already? Do you have entries where one of the pair (forward or reverse) is missing?

[Learner]

@Damian Kao great point, how to check for those ? wow, thanks for such a comment

Answer 1

[h.mon]

To parse fastq files, you may use MAPK original idea of parsing every four lines:

$count++;
if($count eq 4){ ... }
$count = 0;

Although the fastq specs doesn't make mandatory 4 lines (header, sequence, header, qualities) per record, it is so widely adopted it is nearly a standard.

To parse paired end sequencing, just open, parse and close the R1 file, then open, parse and close the R2 file, then output the overall results.

[Learner]

@h.mon can you please direct me to the right (original idea of MAPK) ?

[h.mon]

What do you mean? I pointed you to the original parsing idea, it is the code snippet above.

[Learner]

@h.mon when I click on his name, it takes me to his all questions not exactly his original idea

[h.mon]

His original idea can be found at the post you linked: Counting nucleotide frequency using perl script