Part:BBa_K300078:Experience

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

User Reviews

UNIQ2690c4113be7a617-partinfo-00000000-QINU

•••• UNIPV-Pavia iGEM 2010

This E. coli strain has been successfully made competent following a slightly modified version of the protocol described in [Sambrook J, Fritsch EF, and Maniatis T (1989), Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.], yielding 10^6 CFU/ug of DNA after bacterial transformation of pSB*** series vectors plated on Cm or Amp. Briefly, cells were grown to and OD600 of ~0.4-0.6, harvested (4000 rpm, 10 min, 4°C) and the supernatant discarded. Cells were resuspended in (30 ml for each 50 ml of initial culture) pre-chilled Mg-Ca buffer (80 mM MgCl2, 20 mM CaCl2), centrifuged as before and the supernatant discarded. Cells were resuspended in (2 ml for each 50 ml of initial culture) pre-chilled Ca buffer (100 mM CaCl2, 15% glycerol), aliquoted in 0.5 ml tubes and freezed immediately at -80°C. Test the transformation efficiency in Colony Forming Units (CFU)/ug of transformed DNA.

UNIQ2690c4113be7a617-partinfo-00000002-QINU