Oligonucleotide directed mutagenesis

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wesida
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Oligonucleotide directed mutagenesis

Post by wesida » Sun Mar 30, 2008 11:22 pm

Hello everyone,
I have a question about Oligo-N directed mutagensis. after the primer is set and the DNA(plasmid) is replicated, this Hetroduplex is moved into a bacterium (Ecoli). theoretically when the cells replicate each DNA piece (mutated DNA & WildType DNA) will have equal chance of segregating into the progeny cells. BUT... the Wildtype DNA frequency will be bigger than the mutant frequency. This is i think because the DNA repair system will repair the mutated DNA (unmethylated)
IS this the main reason why the frequency of WT will be higher? and Also what are different techniques to overcome this problem (that is to increase the mutant frequency DNA)
ANOTHER QUESTION
What is the significant of the Long primer during PCR mutagensis?
Thanks
hope i didn't write the question in a confusing way

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