what are the HUGE colonies on my negative plates? - (Oct/19/2006 )

this is really depressing. i guess im giving up cloning. so my ligation mix is plated. plates which supposedly contain my insert (still checking by PCR) have only small colonies. and plates that are negative controls have both small and HUGE colonies on them. there are no huge colonies on the positive plates. this is not the first time it happens. i dont think competant cells or SOC are contaminated. maybe the cuvettes since they are being reused forever. but why only (supposedly) negative plates?

this is really depressing. i guess im giving up cloning. so my ligation mix is plated. plates which supposedly contain my insert (still checking by PCR) have only small colonies. and plates that are negative controls have both small and HUGE colonies on them. there are no huge colonies on the positive plates. this is not the first time it happens. i dont think competant cells or SOC are contaminated. maybe the cuvettes since they are being reused forever. but why only (supposedly) negative plates?

scolix, one plate is just the vector without the insert, and the other one is competant cells and SOC.

just a random though, could the place where you store/make/streak you plates be contaminated?

It is just that when I get this problem of overly large colonies.. it usually is the plate which is contaminated. It might be the case here.

it is true i also thought of this. while i do my plates on the day of electroporation, and dry them in the culture hood....it is still possible...however im confused why is it always only the negative plates!!!???

has it always been the negative plates even on other transformations? It could well be just blind luck that the negative plates were infected in this transformation

kathy i guess we have the same problem, my negative plates also has this very large colonies . Vector and with no insert DNA