RAPD PCR sequencing

Jack Kramer kramer at news.miami.edu
Sat Jul 11 23:26:06 EST 1992


In article <9207111900.AA11603 at umailsrv0.UMD.EDU> William_D_WARREN at UMAIL.UMD.EDU (ww40) writes:
concerning direct PCR sequencing of RAPD fragments:

>Direct sequencing does seem to be a problem but it should be relatively
>easy to excise the band of interest and clone it into a plasmid
>(using either the TA cloning or blunt end ligation) and then use the
>universal priming sites in the plasmid to sequence from with the standard
>methods.

This is the anticipated method.  I was wondering if anyone has found
a direct PCR method using the same primers.  It would be much simpler
and faster.  Especially if there may be very large numbers of fragments
to sequence.

>I am however rather interested to know what type of information
>you expect to get from sequencing RAPD "amplimers" ???

If one finds unique fragments that correlate with specific
characteristics of segregating populations such as resistance
or susceptibility to a particular pathogen if may be interesting
to check the sequence against Genbank.  Similarity to other known
sequences might provide clues to the Genes involved in the host
pathogen compatibility.

Unique fragments might also provide unique diagnostic probe sequences.



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