Unstable clones - (Apr/19/2006 )

Hello, I'm trying to make a plasmid containing a huge hairpin (500bp). When I'm testing my plasmids about 15% of them has deleted te hairpin-structure. Does anyone have any suggestions, like competent cells, etc.?
Thank you very much.

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You might try a recA mutant background...

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Stbl cells from invitrogen or SURE cells (stratagene if I'm not mistaken).

Also: growing your cells @ 30°C instead of 37 with milder shaking (180 rpm) helps...

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Thank you, I've used Stratagenes SURE2 strain and I'm going to test Stbl3 train from Invitrogen. I've also tested the growth at 30 degreed, but although the ratio between 'good' and 'bad' plasmids in the mixture is better that when I grow my preps at 37 degrees.
But I will try the milder shaking.

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