plasmid integrated or not ?
Theo at mendel.Berkeley.EDU
Mon Mar 21 16:38:35 EST 1994
In article <19940317162325.smithwhi at preiss2.bch.msu.edu>,
Brian Smith-White <smithwhi at students.msu.edu> wrote:
>In Article <2m30lr$mi3 at agate.berkeley.edu> "Theo at mendel.Berkeley.EDU (AT)" says:
>> The following question was posted to me by a friend. I told
>> him to use inverse PCR (i.e. shearing the DNA into small fragments
>> -> self ligation -> inverse PCR -> subcloning and sequencing
>> -> designing new primers -> PCR across the junction betwwen
>> the vector and genomic sequence using unsheared genomic DNA
> Why do the second PCR step? If the product of the first PCR is larger
>than the distance between the two primers, what other interpretation could
>you generate? Would the second PCR be able to discriminate between this
>interpretation and integration in the genome?
This is to role out possible artefact created during the ligation
before inverse PCR.
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