help in PCR
(by rijutak from googlemail.com)
Wed Jun 24 12:46:23 EST 2009
> From: KUNAL KALE <biotech.kunal13 from gmail.com>
> Subject: help in PCR
> To: "methods from iubio.bio.indiana.edu" <methods from magpie.bio.indiana.edu>
> <fda120320906240127w4cc5a5cwac064bf1014e57ee from mail.gmail.com>
> Content-Type: text/plain; charset="iso-8859-1"
> I am studant of PhD. in S.G.B. Amravati university , (India). and I am
> Working on Chloroplast Gene. I am working on an amplification of a gene
> is 1369 bp. I have problem in amplification. With this mail I m sending
> attachment of sample of result which obtain after PCR . primers have been
> checked and seem to be working fine . I am Cerrently Using biometra
> gradient thermo cycler . I am cheking other primer of mec A gene which give
> Proper result .pleas give me suggetion on this problem.Your valuble
> suggetion Definatly help me in my research .
> If anyone has experience amplifying fragments this large I would love to
> discuss the methods by which they are using.
> your sincerly
> Mr. kunal kale
Did you vary other PCR conditions, like the primer annealing temperature or
the number of cycles? Did you add additives like BSA to check whether your
desired product is formed? Also, did you try varying the primer
concentration in your PCR mix? While amplifying such a big fragment, the
choice of polymerase is also very crucial.
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