Contributors: Tank
Received Project A and Project B mutant sequences back from sequencing. Next planned step: MUG assay of the mutants that survived cellobiose selection 2, to test for beta-glucosidase activity.
BL21 re-transformation with CxnA construct
Previous attempt formed lawns, so concentration was lowered this round: 100 µl BL21 competent cells + 2 µl miniprep DNA, vortex, 30 min on ice, heat shock 42 °C for 40s, ~30s on ice, 1 mL SOC, 1h at 37 °C shaking. Plated 25 µl and 50 µl onto separate LB + chloramphenicol plates; remaining cells spun down, resuspended in 100 µl SOC, and plated concentrated onto a third plate. Incubated overnight.
M9 control plates
4 Duran bottles prepared (62.5 mL 4X M9 + 157.5 mL water + 7.5 g agar each), autoclaved.
Result: plates came out brown — consistent with the contamination/Maillard-type concern flagged on 6 July.