Colony PCRs were performed on each of the transformations from 20151015 (RLOv_ DNA_helicase, RLOv_head_to_tail, RLOv_membrane_gene_1, RLOv_membrane_gene_2, RLOv_tail_to_fiber) to confirm successful ligations in plasmid pCR2.1 using the M13F/R vector primers.
Colonies were picked form the transformation plates with pipette tips, re-streaked on a secondary, gridded, numbered LBAmp100+x-gal plate and then used to inoculate the respective PCR reactions.
Six white colonies (positive clones) and a single blue colony (negative clone) were selected from each transformation.
Master mix calcs are here (Google Sheet): 20151019 – Colony PCRs RLOv
Restreaked plates were incubated @ 37C O/N and then stored @ 4C (Parafilmed).
30μL of each reaction was run on a 1% agarose 1x Low TAE gel, stained w/EtBr.
All the PCRs look good. All white colonies selected contain a PCR product of appropriate size (i.e. larger than the blue colonies; negative [-C] control). Will select clones #1 from each to grow up for plasmid prep.