On Sat, Mar 10, 2012 at 2:03 PM, Jeswin <phillyj101@gmail.com> wrote:
> On Sat, Mar 10, 2012 at 1:05 PM, Nathan McCorkle <nmz787@gmail.com> wrote:
>> Jm109 or DH5-alpha or K12... whatever you have on hand.... i did this
>> once (with a commercial electroporator) and it worked wayyyy better
>> than CaCl2
>>
> Where do you get those from?
NEB, Life Tech, etc... basically any lab strain e.coli used for
cloning or expression is based on those strains. Historically I think
it began with removing virulent stuff, then making certain mutations.
>
> The ones I use are New England Biolabs Competent E coli. Are they
> special strains that have improved transformation efficiency over the
> DH5-alpha, etc?
The JM series were made for cloning in the 70's or 80's, and it turns
out NEB actually carries a few of them:
http://www.neb.com/nebecomm/tech_reference/restriction_enzymes/ecoli/strains.pdf
The mutations for efficiency are something you'll have to dig into
yourself, but all the strains have genotypes along with them for
comparison, but yeah generally cloning strains have some mutations to
make foreign DNA less prone to being rejected.
>
>> just make sure its in exponential phase growth, that's all
>>
>> I rinsed my e. coli 2 or 3 times with pure water before adding DNA and
>> electroporating, but Mega says he doesn't have a centrifuge... and if
>
> What do you mean by rinsing with water?
From my LB liquid culture I centrifuged, decanted supernatant,
resuspended in distilled/sterile/nanopure water, repeat once or twice
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--
Nathan McCorkle
Rochester Institute of Technology
College of Science, Biotechnology/Bioinformatics
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