About reporting a new splice variant in NCBI - (Aug/18/2009 )

Hi!!

I believe I have identified a new exon in my gene of interest using RT-PCR. Until now there are 3 confirmed exons.
I'm using a primer pair that anneals with exon no2 and no3. These two exons are parted by a 22000bp intron.
Apart from the expected band, I was getting a larger band which turned out to be a possible different exon. The sequencing result showed that the larger band has the expected sequence + 120bp from the 22000 intron in between.

So my question is, what has to be done now (experimentally) in order to have valid proof that there is a different splice variant and submit the new data in NCBI.

I'm thinking as a first step of scanning the whole mrna using primers that anneal at the beginning and at the end of the known transcript in order to check for additional changes.

Thanks in advance!!

haris on Aug 18 2009, 01:16 PM said:

Nothing. If you have amplified a cDNA and have determined the sequence of this cDNA, you can submit it to GenBank like that. Of course, if you want to publish your work describing a new isoform of your GOI in a journal, you do need to do some more experiments. One exp you could do is to detect in which cell lines/tissues you find the new exon. Additionally, does the new exon encode any protein domains that aren't present in the other transcripts? It all depends what you want to do with it.