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CD6 T-DNA lines

Carolyn Schultz c.schultz at botany.unimelb.edu.au
Thu Jul 16 11:07:16 EST 1998

I have been screening the Feldman T-DNA lines for an insert in a
specific gene using a PCR technique.  Using gene specific left and right
primers in combination with T-DNA border primers I have screened the CD5
pools without any problems.  However when attempting to screen the CD6
pools I have not managed to amplify either the actin control or controls
using the left and right primers for the gene on the pools of DNA.  My
conditions are: 94.0 C for 20seconds, 55.0 C for 30seconds, 72.0 C for 1
minute, x 40 cycles.  My concentration of MgCl is 2mM, primers, approx
100pmol, and DNA using both 25ng/ul and 8.3ng/ul.  Any advice on how to
optimise my conditions or information on problems that others have also
encounted using these pools would be greatly appreciated.  Look forward
to hearing any responses,

Kim Johnson
Plant Sciences and Biotechnology Centre
University of Melbourne.

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