Number of integrations in transgenic plant

a_firzinger at my-deja.com a_firzinger at my-deja.com
Fri Oct 29 05:03:51 EST 1999


We got a transgenic plant and know the sequence of the
integrated construct. However, we do not know how often
the construct was integrated, which will be essential
for further analyses. We can isolate DNA, make PCRs
and do sequencing with our equipment, but we cannot clone or
grow bacterias or plants. And since
Southerns are quite labourious and we would have to establish
nearly everything new for Southern blotting, we are looking
for other methods easier to handle with our resources.
There are some methods available to get the sequence of
unknown flanking DNA of integrated known sequences just
by utilizing only PCR and isolation of resulting fragments,
(Laskus et al.(1999).Detection and sequence analysis of HepatitisB
Virus integration in peripheral blood mononuclear cells;
J Virol 1999; 73:1235-1238)

My question is, if this methods are applicable if the
construct integrated more than one time in the genome ?
Or does someone else have a protocol for checking the
number of integrations in a nearly unknown genome, just
by using PCR.

regards

Andreas Firzinger


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