selective splicing? - selective splicing? (Oct/11/2008 )
I have done RT-PCR of my gene of interest in arabidopsis, but after running the gel ,there are three different size of bands. I cut each band in the gel and sequenced them, unfortunately, the signal is complex.
i donot know the reason, but some inclues indicate that this gene may have selective splicing, but not sure, needing sequencing result to validate.
others suggest me to use the cloning method to sequence different RT-PCR product.
and any other good suggestion? about my gene (a PPR domain protein) or about my experiment?
thank you!
i donot know the reason, but some inclues indicate that this gene may have selective splicing, but not sure, needing sequencing result to validate.
others suggest me to use the cloning method to sequence different RT-PCR product.
and any other good suggestion? about my gene (a PPR domain protein) or about my experiment?
thank you!
Hi Kingber
I also found that some genes perhaps take selective splicing. and you can search the accession number in TAIR which will tell you whether it has seletive splicing.
Stone
thank you! stone
I have checked the tair, no selective splicing.but I sequenced a fragment of this gene, finding that the prediction of tair is not very correct about the CDS.
I have checked the tair, no selective splicing.but I sequenced a fragment of this gene, finding that the prediction of tair is not very correct about the CDS.
yes perhaps you need to choose sequence view button. And that will be more exact.
stone
I think you should clone your DNA once you have gel extracted them.
This way, you will be sure that you are sequencing a single DNA fragment at a time and avoid any strange results.
use a TOPO cloning kit, it's straightforward.
and what about trying to prepare your cDNA with your gene-specific primers, it might give more insight to your problem if you do not obtain 3 bands this way as well.