Please Help: human Sox2, Oct 4 promoter region cloning
Posted 16 January 2010 - 08:53 PM
I am a new guy working on Stem cell filed. I try to clone Human Sox2, Oc4 promoter region into pEGFP-reporter vectors. I am a little confused about two points:
1. Which type of cells genomic DNA could be use to amplify the upstream of Sox2 or Oct4 genes? Could I use the normal 293T cells or Hela cells to amplify the promoter region? Or Should I use any human Stem cells or IPS cells? if so, which type of cells should I use?
2. Where could I find the promoter region sequence? Or I only clone 2 kb of the upstream of the genes?
Any one has adviec? Thanks a lot in advance
Posted 17 January 2010 - 07:41 PM
293T cells are okay for your amplifying the promoter region. Try to avoid use abnormal cells such as tumor cells to amplify due to the gene in that cells may be abnormal. But for cDNA clone ,you have to use stem cells.
For promoter region sequence, you'd better look up for some literature concerning Sox2 or Oct4 promoters. If there is none, you can use http://www-bimas.cit.../molbio/proscan to predict. Try to clone a longer region in case not missing any information.
Posted 18 January 2010 - 08:45 AM