Frequently Asked Questions about Common Vectors
Antibiotic Usage
Transformations
Chloramphenicol - what concentration should I use?
Chloramphenicol concentration requirements in plates versus media is different. At the DGRC, we make a 34 mg/ml stock solution in 100% ethanol (store stock at -20). Then for liquid cultures, add 2 microliters of stock to for every 1 ml of LB liquid culture (you can also get by with 1 microliter per ml if necessary). For plates, add 1 ml of stock chloramphenicol solution for every 1 Liter of media for plates.
Can I use electorporation on the Whatman discs?
Yes - Whatman has documented that this does work. We have not tried it at the DGRC but we have received the following information from a colleague who succesfully used electroporation: After rinsing with the TE for a few seconds he added electrocompetent XLI Blue cells (the lab's homemade) to the disc and incubated on ice for one minute. He then transferred the cells to a cuvette and electroporated. He plated the entire transformation on amp plates and obtained around 5 x 10^3 colonies.
Can I use glycogen precipitation?
We know of one person who tried a glycogen precipitation and centrifugation protocol as an alternative for recovering DNA from discs and did not have great success. We do not recommend trying this procedure as a means of recovering.
