Hello,

I was making DH5a competent cells but I was confused with the condition of bacterial culture in the protocol I followed.

The second step of the protocol is to "Dilute 20 μl of overnight culture into 10 ml of LB broth. Grow at 37◦C with shaking to early log phase (OD600 = 0.3)", which means the dilution ratio of the culture is 1:500. I'm wondering how long should it take to grow a bacterial culture with 1:500 dilution ratio to early log phase? I followed the protocol and it took me almost 5 hours to grow the culture. Is it normal to take really long time like this?

Is the 1:500 dilution an error in the protocol?(maybe it should be diluting 200μl of overnight culture instead of 20μl?) Or this is for particular purpose and I should follow the protocol?

Thanks for answering!