RAPD PCR sequencing

ww40 William_D_WARREN at UMAIL.UMD.EDU
Mon Jul 13 22:53:00 EST 1992


>>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.
>

Jack,
I'm glad it you and not me !!! I collegue of mine calculated that,
in Drosophila, it would take at least 30 cycles of repeated outcrossings
to replace the genetic background to within approximately 5 kb of
the allele of interest. Clearly this figure would be different depending
on cross-over frequency and genome size in "Your" organism. So it
seems to me that even if you are extremely rigorous in checking that a
particular RAPD cannot be separated by crossing-over from the trait of
interest, your amplimer sequences may well be several kb, if not 10's or 100's
of kb from the gene you are trying to get at. Genes whose function in
the vast majority of cases you have absolutely no idea about.

Good luck (I think you're goin' to need it :)

Bill Warren
Ctr. Ag. Biotech.
Univ. of Maryland





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