I have some crispr mutants that have indel mutations, mostly 1bp insertion or deletion. Most 1bp indel resulted in pre-mature stop codons during translation. However, I don't know whether the mRNA level of genes having indel mutations were affected and why.
Thank you,
Yuan
Hi Yuan,
if the modifications occurred in coding sequence there is no clue that transcription would change, but it's not the case if those modifications touch the 5' or 3' UTR where most of the regulation sequences are lying. the only changes will be in the transcripts due to frameshifts.
fred
It would depend upon whether you are referring to prokaryotic or eukaryotic organisms. In prokaryotes you have transcription and translation frequently coupled, so an in-del mutation (ie a frameshift) would cause the cessation of translation, and if this occurred fairly early in a gene (ie first half or so) then you are likely to have a negative effect on transcription. This is essentially the explanation of polar mutation in bacteria. This would not be the case in eukaryotes.
Hi Yuan,
I don't know that the indel would affect the rate of transcription per se, but most indels will typically result in premature stop codons. It is likely that the mRNA would then be degraded by nonsense-mediated decay. Therefore, if you were to measure transcript abundance by qRT-PCR it is likely you would see a reduction compared to wild-type.
Josh
Thank you so much for all the answers. From Joshua Clayton answer, I learned what is nonsense-mediated decay. So I will use qRT-PCR to check transcription level of my gene with indel mutations.
I use Arabidopsis (a model plant species) to create crispr mutants.
Frederic Lepretre and Michael J. Benedik
Why do you think the indel mutations won't affect transcription? By the way, most of the mutations I have resulted in pre-mature stop codons.
Thank you again.
Yuan
Yuan - in bacteria transcription and translation are coupled so translation can have an effect on transcription. An indel creating a nonsense mutation would not have an effect on the rate of transcription in a eukaryote, but it might in a prokaryote. However Joshua is correct that nonsense mediated decay could have an effect on turnover of the mRNA and hence reduce the overall level of a transcript (without affecting its rate of transcription).
Michael J. Benedik Thank you for your answer. I will go ahead check their transcription level.