filter illumina index

Question

[priscilladraj]

Hi, I ran an iseq illumina sequencing on my libary. And due some sample prepration problem, I have a low concentration of my libary. Hence, after sequecing and base calling, the iseq base caller grouped all my reads (dual index, 3 pooled libary = 6 reads total) under a undetermined folder. I want to know how can i still demultiplex my data that is in the undeterrminded folder? I tried using the https://pypi.org/project/filter-illumina-index/ script. BUt still cant or have I made any mistakes in my code

filter_illumina_index --1.0.4 -f FILTERED -u UNFILTERED  -i ATTACTCG -m 0 -t 1 -l 6 -v 0 /home/ubuntu/dnaread1/20200903_FS10000764_45_BPC29622-2333/Alignment_1/20200904_164251/Fastq/Undetermined_S0_L001_R1_001.fastq.gz

I thought I should demultiplex each index one by one then I can filter out my data for further analysis.

Could someone help me please. I am super new still to any demultiplxing any data

Pris

[Gabriel R.]

Just so I can understand, could you post examples of sequences you have and a list of expected indices?

Answer 1

[priscilladraj]

thanks I manages to resolve it demultiplexing it again bcl base caller software and it worked this time because I learned that teh Iseq reads i5 from reverse complement direction. So its my mistake of not knowing this.

Answer 2

[GenoMax]

You should demultiplex data based on solution I show here: C: demuxbyname.sh output help

There may be other unexpected index combinations present. That is normal to some extent (< 5% of total) and those can be ignored.