Same exon-intron junction sequences for all genes? - (May/10/2005 )
I wonder do the genes have the same exon-intron junction sequences or do they have different junction sequences? I was told that all genes have this general junction sequences of the exon-intron-exon:
The arrows indicate the borders/junctions of exons and intron. When this intron is cut out and the exons are united we would have this exon-exon sequence:
How can i check which sequences are exon-intron junctions and which are not?
Not all exon intron boundaries are identical but I think you have the general idea. Unfortunately not all sites that conform to this rule are used and not all sites comply strictly to this rule. This means that you can't just search for consensus splice sites and assume they are real. The easiest way to find intron/exon boundaries (designing PCR primers?) is to align cDND and genomic sequences. If its in the genomic and not the cDNA it is an intron. (be careful at 5' ends however as not all cDNA sequences are complete and should be avoided for PCR because of this).
Yes, it is for primer design. Do you know why the 5' cDNA end is not complete? So when i design primers then they should be in the middle of cDNA for best result?
Thanks for your great reply.
When the reverse transcription reaction proceeds the reverse transcriptase will "drop off" the RNA template leading to incomplete extension products which are missing the 5' end. For best results its best to have the primers near the 3' end so that if you do get an incomplete reverse transcription it will not effect your PCR. I will add also that the termination point at which transcription finishes can be variable so to be safe stay in the coding sequence and not at the extreme 3' end either!
Thanks alot for your help!