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Germline transform by electroporation

Chris Merli Chris_Merli at qms1.life.uiuc.edu
Wed May 26 12:45:11 EST 1993


I am attempting to do germline transformations of D. melanagaster using
electroporation.  I have been following the basic protocol of Kim Kamdar
[NAR 20:(13) pp352; pers. comm.] but I noticed low survivability in
non-dechorinated embryos at 250v/cm and 960uf.  In short I collect 100-1000
embryos, 0-1 hr old, suspend them in 800ul of 10ug/ml plasmid + 2ug/ml
transposase plasmid + 5mm KCl + 0.1mm sodium phosphate pH 7.8 at room
temperature.  I then pulse twice in a Bio-Rad electroporation apparatus,
wait two minutes, drain, and place in fly bottles.  I get less than 20%
survivers under these conditions but I have not yet finished the crosses to
determine the number of transformants.  I would like to know if anyone else
is attempting this method and what success they are having.

Chris Merli



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