hi...
i am doing research in cancer. I bought new primers for my work.Tm for tese forwrd and reverse primers are 73.3 and 72.9 respectively. I did PCR with these. but i didnt get any product in any temperature(45-72c).pleare give some suggestions.
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#2
GNANA
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Posted 05 May 2011 - 03:07 AM
if you hav no other option than this primer you can very well try to optimise the condition for this primer itself, but will demand time and tricky....fine what is your template???is it DNA, plasmid or cDNA??? What PCR mix you are using, is it a readymade mastermix or the conventional tag+dntps+mgcl2 etc??? what abt the GC content of your region to be amplified??? if gDNA how pure is your template??? so all these things matters....so answer for all these queries you will find some ways to troubleshoot your pcr in this forum...
good luck,
GNANA...,
good luck,
GNANA...,
I would prefer being perfectionist rather than a passionist in Research.
I always had an alternate hypothesis....
I always had an alternate hypothesis....
#3
Adrian K
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Posted 05 May 2011 - 08:10 PM
Just curious: when you do your gradient from 45-72C, how many temperature difference between each interval? 45C, 46C, 47C, 48C..... or 45C, 50C, 55C....?
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..."best of our knowledge, as far as we know this had never been reported before, though I can't possible read all the published journals on earth, but by perform thorough search in google, the keywords did not match any documents"...
"what doesn't kill you, makes you stronger"---Goddess Casandra reminds me to be strong
"It's all just DNA. Do it."---phage434
..."best of our knowledge, as far as we know this had never been reported before, though I can't possible read all the published journals on earth, but by perform thorough search in google, the keywords did not match any documents"...
"what doesn't kill you, makes you stronger"---Goddess Casandra reminds me to be strong
"It's all just DNA. Do it."---phage434
