E. coli Strains

About microscopic forms of life, including Bacteria, Archea, protozoans, algae and fungi. Topics relating to viruses, viroids and prions also belong here.

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nikinje
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E. coli Strains

Post by nikinje » Fri Feb 13, 2009 5:55 pm

This is a follow up to my dead E. coli problem (posted already on the forum).

I have been using a kit from Clontech called in-fusion 2.0 for cloning, produced my clones and transformed them into their "super competent" FusionBlue E. coli cells. This worked fine and produced many colonies, but I am keen to get back to using a more common generic cloning strain used in our lab already, or common in other labs.

I have tried to transform the same clones into DH5 alpha cells, but no colonies grew. For these reactions you need "super competent" cells (according to the website), with a transformation efficiency of greater than 10^8 cfu/ug DNA (that doesn't mean anything to me, but if it does to anyone else?....).

I was wondering if anyone could suggest any strains I could try, I've been told to try Invitrogen's Top10 E. coli or Excel-1-Blue. Are there any others which could be useful?

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canalon
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Post by canalon » Fri Feb 13, 2009 7:21 pm

"10^8 cfu/ug DNA" means that for each µg of DNA in your transformation mix, you will obtain 10^8 cfu (colony forming units, in this case: viable transformants).

I cannot suggest any special strains but many of the commercially available strains are not the healthiest possible bacteria as they have had a few genes knocked out to improve the transformation efficiency. Plus the buffers and solutions in which they are stored ready to use have been developed to improve efficiency, that explains why kits usually gives better yield than in-house bacterial cultures. However, if the quantity of transforming DNA can be increased, this can compensate for lesser efficiency
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