I am using Roche 3'/5' 2nd generation RACE kit to determine 3' and 5' utr regions. I have completed all steps and sequenced my 3' plasmids (pGEM-T Easy vector).
The main point: my gene has a transmembrane domain including stop codon (approximately 90bp). My sequence results shows that my sequence finished just 2 bases after transmembrane domain.
My sequence contains:
Starts with my primer sequence and then my sequence except last 90bp which is coding transmembrane domain; after that polyA (AAAAAAAA) and the rest is pGEM-T vector sequences.
is it possible that you have a alternative transcript (e.g.. for a soluble variant of the protein), without the transmembrane domain?are there potential splicing/donor acceptor sites flanking your 90 bp? (often GT/AG)
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