cDNA cloning by PCR
Posted 17 March 2005 - 02:43 AM
i have purified a protein whose N-terminal sequence is known. If sequence of only Nterminal is known how shuould i design the primers to get the amplifcation of the desired gene can i get a cDNA clone from a single primer .what should be the primer for other flanking region of the gene?
Posted 17 March 2005 - 11:44 AM
Can I know from which organism that you isolated your protein?
How many base pair that you know?
I need to know those information before I can know what to do.
Posted 17 March 2005 - 10:32 PM
Posted 29 March 2005 - 02:18 AM
To get the primer for other flanking region of the gene, I think is good to use bioinformatic software and make use of swissprot protein database.
Now you know 15aa (=45bp). load your know 45bp on NCBI and BLAST. From the BLAST search, the programe will show you a lot of similar protein. You can make use the information from the data base predict your reverse primer.