The results of my big transformation experiment are clear but not what I had hoped.
There's no evidence of transformation of either strain by any of the markers. As expected, the C600 strain produced Lac+ revertants (frequency 10^5 - 10^6) and Leu+ revertants (frequency 10^6 - 10^7). I know these are revertants because the control cells (no DNA or no sxy insert or no inducer) produced just as many Lac+ and Leu+ colonies as the cells given DNA, sxy and inducer. The thr-1 mutation didn't revert detectably (frequency <10^8) style="font-style: italic;">lacZ gene.
I know the plates used for the selection were OK because the donor strain (w3110) grows fine on all of them but the recipients don't. This also confirms that the donor strain carries the expected wildtype alleles of lac, leu and thr.
So what next? First the RA and I need to discuss her latest results checking out the crp::kan marker she's been using for her (apparently successful) transformations. Then I'll probably do another set of transformations and controls, this time with that marker.
Joys of gardening - late fall edition
8 hours ago in The Phytophactor