HELP! Desperately need a dot blot method
greggsilk at aol.com
Tue Jun 30 07:19:06 EST 1998
>I may have a simple solution to your problem. When I do colony lifts of
>E.coli containing plasmids of interest I use the "Turbo Juice" method.
>Basically this involves using dry membrane to lift the colonies. The
>membrane is then put DNA side up on a bit of 3M paper that has been
>soaked with Turbo Juice (simply 2x SSC + 5% SDS) and popped in the
>microwave on high for 3-4 minutes until very hot and dry. The membrane
>can then be hybridised straight away; no washing or UV linking required,
>just pop straight into prehyb then hyb mix.
>Anyway, often when I do lifts like this I just spot my positive and
>negative control plasmids on an offcut of dry membrane and pop then
>through the same pocedure as above after the spots have dried a few
>minutes. I know it seams to easy to be true but it always works. No
>reason why it should not work for a whole blot of spotted plasmids.
I do something similar- lift colonies with dry Nytran, layer between 3MM,
autoclave 3 minutes, then prehyb. 'Works like a champ.
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