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TOP10 chemically competent cells: Difference between revisions
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This protocol is a variant of the Hanahan protocol using CCMB80 buffer for DH10B and TOP10 strains. It builds on Example 2 of the [[Media:pat6855494.pdf | Bloom05 patent]] as well. | This protocol is a variant of the Hanahan protocol <cite>Hanahan91</cite> using CCMB80 buffer for DH10B and TOP10 strains. It builds on Example 2 of the [[Media:pat6855494.pdf | Bloom05 patent]] as well. | ||
===Preparing glassware and media=== | ===Preparing glassware and media=== | ||
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* Plate 20 μl on AMP plates using 3.5 mm glass beads | * Plate 20 μl on AMP plates using 3.5 mm glass beads | ||
* Transformation efficiency is 15 x colony count x 10<sup>5</sup> | * Transformation efficiency is 15 x colony count x 10<sup>5</sup> | ||
==References== | |||
<biblio> | |||
# Hanahan91 pmid=1943786 | |||
# Bloom05 US Patent 6,855,494 [[Media:pat6855494.pdf]] | |||
</biblio> | |||
6/7/06 colony counts: | 6/7/06 colony counts: | ||
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15/ cells are grown in 15/10 medium. Slow growth, eliminate this idea. | 15/ cells are grown in 15/10 medium. Slow growth, eliminate this idea. | ||
Dispose of M, 15/1, 15/2, 15/3 cells. Redoing competent cells today. | Dispose of M, 15/1, 15/2, 15/3 cells. Redoing competent cells today. | ||
m/s1/s2/s3 cells are successive versions of competent cells grown in SOB medium. | |||
