C. T. Chung published a protocol for one-step preparation of #competentcells in April,1989. This has been one of the easiest protocol.
FIGURE: Centrifuged transformed cells in LB just before plating!
Recipe of 2X #TSSbuffer:
1. 10% (w/v) Polyethylene Glycol-8000 or 6000 (#PEG8000 or 6000), the original article suggested PEG-3350 can also be used.
2. 5% (v/v) Dimethyl Sulfoxide (#DMSO)
3. 50mM Mg2+ (#MgSO4 or #MgCl2), the original article suggested 20-50mM Mg2+ salts.
4. These components are dissolved in #LuriaBroth and pH is adjusted to 6.5.
Protocol:
1. An overnight #culture of the E. coli cells are diluted 1:50 and incubated at 37C and 200-225RPM until the #OD at 600nm reaches 0.3 to 0.4 (early log phase).
2. An equal volume of ice cold 2X TSS buffer is then added to the culture, and 100ul of this mix is then aliquoted in previously freezed 1.5ml tubes over ice.
3. For long term storage, place the aliquots in -80C. The transformation efficiency of the cells are at least 10^7 transformants per ug of #DNA used, and this is maintained in -80C for 1-18 weeks.
Uses
To prepare E. coli competent cells to be subsequently used for transformation.
More to munch on: