PCR and competent cells - (Jun/13/2008 )

Hmm...just wondering if its possible to get a product from a no DNA sample in PCR even though you have added in the other components correctly...

And is there a difference in using overnight cultures rather than those at log phase when preparing competent cells? Thanks for the heads up guys!

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Re-PCR... if you havent added template and still get a product, check all your reagents because one of them is contaminated. Usually is the water or the primers. Prepare your reactions with all fresh reagents, and if you really want to know which is the dodgy one, all fresh reagents but one (check each of the old ones).

Re-competent cells: I'm not exactly sure of the explanation, but my guess is that there'll definitely be a difference, I've always used cells in the exponential growth phase to make competents.

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1. You might have what we in the PCR 'biz' call contamination. Clean all pipettes, change over your reagents, and set up in clean air. This is especially important if your target is commonly used.
2. Overnight cultures are in the vegetative step, and make for poor to useless cells for transfections. You can make an overnight, but you must grow up fresh on the day you prepare the cells.

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swanny is right,

just want to add a lil stuff. Please use filter tips. Somehow the aerosol has something to do with this I guess. Just use filter tips for PCR.

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