We’ve had a disabled cart sitting in lab for months that has locked up wheels and all of the bolts attaching the wheels to the cart are so rusted, the bolts and nuts cannot be separated by normal means (believe me, I’ve tried numerous methods over the last few months to no avail). Since we’re planning a significant lab cleanup next week, having this cart repaired will be good, so that it’s not sitting upside down in the lab any more AND fixing it will provide us with a second cart for cleanup day!
In any case, I ended up having to drill out the rusted bolts/nuts on three of the four wheels. After that, I put on the new wheel hardware and it’s good as new!
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.
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.