directional cloning problem!

chengzhjun at chengzhjun at
Fri Aug 23 04:54:15 EST 2002

recently i performed directional cloning but can't make it. so i want to know where the problem is. at first i made my pcr product cut by NdeI and XhoI,then the plasmid i used is pET28b cut by the same two enzyme. secondly the mixture of pcr product and the plasmid after enzyme cut ligated with the T4 ligase.At last i can't get the transformant on the plate.
  so where is perhaps the main problem during the procedure? please kindly offer your suggestion! thanks .


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