Hi,Can I use a 2.6kb gene (whole gene instead of mini-gene)...

Aditya Agarwal
Aditya Agarwal
Sep 23, 2021
protocol Protocol: Assaying the Effects of Splice Site Variants by Exon Trapping in a Mammalian Cell Line
Hi,
Can I use a 2.6kb gene (whole gene instead of mini-gene) as an insert in this vector?

Thanks!
Aditya
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Stuart W J Tompson Answered Sep 26, 2021

University of Wisconsin-Madison

I am very doubtful that this would work. Why would you want to do this experiment? This assay is to test for the presence of exons within a genomic interval or to test whether variants in any known splice site regions could be interrupting the correct recognition of an exon, leading to aberrant splicing. The assay requires that the "experimental exons" that you are testing harbor both splice donor and splice acceptor signals, so that they can be correctly spliced in conjunction with the preceding and following "vector exons". I presume that by "whole 2.6kb gene", you mean a complete cDNA sequence (coding only, i.e. a transcript sequence with all introns removed), in which case by definition all of the splicing signals have already been removed from your sequence via splicing of the mRNA. Hope that answers your question!
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