High Throughput transformation - (Jun/27/2007 )

Hello every one


I need help I want to do transformation by High Throughput

I did it by this process:

Thawing 10 min the DH50-alpha
Add the ligation mix
Incubation 30 min on ice
Heat 2 min in 42c
Shacking 1hr at 37c 225 rpm
Plating on Petri dish for O/N

The problem most of my Petri dish did not have any colony this is the problem
So is there any protocol for High Throughput transformation


I hope every one understand me

how much ligation mix do you add to the competent cell???
my heat shock is for 30 sec and after i shake it in 150 rpm, i think 250 rpm is too much

Hi,
How's the control? Transformation efficiency of the cell?

i added 3ul from the ligation mix


Transformation efficiency of the cell is good

What's the antibiotic-resistance in the plasmid? If it's ampicillin, you shouldn't heat shock the cells. Try this instead:

Pre-heat the plates to 37c for at least 30 minutes.
Thawing 10 min the DH50-alpha
Add the ligation mix
Plating on Petri dish for O/N

my resistance is kanamycin