Wed, Oct 06 21

Gibson assembly and E. coli transformations #

Picked up plates from Gibson assembly reactions yesterday and all samples had at least some colonies which was nice to see. Tada’s VR-30 transformations does not unfortunately.

Given that transformations of VR 8, 12 and 15 were successful I preformed an additional Gibson assembly reaction with the remaining samples (VR 16, 17, 20, 21, 23, 25, 27, and 31) using the reagents and volumes described in table 10-5-21 of the Gibson assembly spreadsheet. After incubating samples at 50C for 1 hour I placed samples into the deli fridge for 5 hours while I helped Tadas with culturing colonies and did class work. At 1:20 PM I transformed electro-compentent cells following previously used NEB protocol with 4 ul of Gibson assembly reaction per sample. At 3:10 I retrieved samples from the 37C room after shaking and plated 200 ul of each culture onto LB + Amp plates which I then places in the 37C hot room to grow overnight.