Hello everyone, Is it mandatory to have same mutant sequence in F1 generation fish to cross between them? how can I seperate fish using sequence data?

09 September 2018 1 5K Report

I have got mutant fishes with CRISPR-cas9 system, and my fishes are now in F1 generation. I have sequenced the mutant region. But I am facing problem to find out the number of base deleted or added. How can I do that using my sequencing data? Moreover, do I need to confirm the same F1 mutant fish(same mutant sequence) before crossing to get F2 fish?

Frederic Lepretre

Hi Ramendu

you should run a blast with the part of the sequence you expect to be mutated against the wild type sequence. bast is a free tool from NCBI and you can use it at (https://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastn&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome). just take care to use the good species and remember that CRISPR can mutate your targeted sequence but due to its inconstancy, other part of the genome can be touched (i.e. off targets).

fred

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