I am planning to perform sodium bisulphite sequencing. Currently, I am trying to design primers of the promoter region of my target genes. However, some of these promoters have not been well-characterized and the exact promoter sequence cannot be directly obtained from PubMed. Therefore, I used a programme, PROMOTER SCAN, to predict the promoter by pasting the 5'upstream sequence from the start codon. But the software said there were no promoters. Whats wrong with that?
How do you guys predict the promoter sequence?
-Samantha-
What organism are you working on? Human?
If it is human, you have pretty good chances to get the predicted promoter sequence from genome databases if there is no experimentally characterized promoter deposited in GenBank. You can go to ensembl.org to search for your gene and then export the 5'-flanking region which is highly likely the 5' regulatory region of your gene. The genome database has annotated human genome sequences using mulitple pipelines. Usually the prediction of 5'-flanking region or promoter is more accurate than stand-alone promoter prediction programs. For example, the pipelines align ESTs to genomic sequences. If a region aligns with many ESTs, this region may not be a non-coding seequence. Further, if this region is located immediately upstream a gene, most likely it is a promoter region. Because you wanna do methylation study on the promoter, the presence of CpG island is very important. After obtaining the sequence, analyze it for CpG islands, if there are, further suggesting the sequence Does contain a promoter.
-pcrman-
thanks for your reply.
actually, i am working on rat genomes.
-Samantha-
I have found a nice website, that will show the genes exons & introns...
You may use the friendly interface to fine the potential promoter...
check it :
-Ming-
