pcr template amount - (May/27/2008 )
I am doing standard pcr. My template is a miniprepped plasmid. I am using pfx dna polymerase from invitrogen. In the begining, how much tempalte should I add? Please give me a range. I checked, some website said 5ng is ok, while some said don't use more than 1ng. I plan to try 10pg and 100pg first. (My final product is 3.6kb).
-hais-
QUOTE (hais @ May 27 2008, 04:29 PM)
I am doing standard pcr. My template is a miniprepped plasmid. I am using pfx dna polymerase from invitrogen. In the begining, how much tempalte should I add? Please give me a range. I checked, some website said 5ng is ok, while some said don't use more than 1ng. I plan to try 10pg and 100pg first. (My final product is 3.6kb).
10pg and 100pg sounds just right. If PCR of your plasmid prep doesn't work, try linearlized plasmid too.
-cellcounter-
A range of concentrations is a good place to start. One of them should work but if none do then come back here because your problem may not be related to the amount of template you've used.
-killerkoz17-