No colonies after LR recombination

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RAD's picture
No colonies after LR recombination

Hello. iam really new to molecular biology. recently, I performed the LR reaction but got no colonies of E.coli. I tried using a new batch of clonase II, leaving the reaction overnight, it didnt work. any suggestions are highly welcome..thanks in advance!

Chin Fen Teo
Chin Fen Teo's picture
 Hi RAD,

 Hi RAD,

Have you confirmed whether you are plating the transformed E. coli on the correct antibiotic containing plate? Almost all of the destination constructs that I work with carry Amp resistant gene.

Also, be sure you follow the transformation protocol "precisely" come along with the invitrogen kit- for instance, the incubation/heat shock time, about thawing the competent cells on ice, about not pipetting the competent cells up and down etc.

Good luck.