Hello,
I have demultiplexed my short reads and I have removed the Illumina adapter left in the sequences and now I need to remove the barcodes used by the sequencing centre from my reads. I have looked into Trimmomatic, Cutadapt (that I have used for the adapter removal) and FLEXBAR. As I have a series of barcodes used throughout my samples (720 samples), FLEXBAR seemed the only one that accepts a .fasta file with the barcodes specified. My question is if I can run the program to detect and remove the barcode sequences across my samples without going through the demultiplexing again.
Thanks
In my opinion if you use cutadapt, you can mention primer sequence. With that it will remove everything left and right from the primer sequences leaving just the variable region of amplicons.
And I guess, there should be no problem of with demultiplexed samples, as cutadapt is most commonly used for demultiplexed samples.
Do you need further help in using cutadapt, please let me know.
Abhijeet Singh Hi and thanks for your reply. I see. This would solve the issue very quickly. But I guess it only works when the both barcodes and adapters are at the same end of the read ? In my case, as my short reads are generated through GBS, the sequence is:
Barcode - DNA sample - Illumina adapter
So the Barcode wouldnt be cut when using cutadapt :(
This is not an unusual case and it can be done easily, and in many different ways. One way can be remove the barcode by fixed number base trimming on left end and adapter trimming with an input adapter file or with fixed number trimming o right end too since the length of both are constant.
If you have not gone through the user manual of cutadapt, I would highly recommend you to do so. Cutadapt have a very2 good documentation page.
https://cutadapt.readthedocs.io/en/stable/guide.html
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