1 reply to this topic

[aravinthkumarj]

Hi,
I cloned 1.4kb insert DNA into my 5.7kb vector. I got colonies and my colony PCR also showed positive results.
But when I digested the cloned plasmid, my result was strange. I got my insert (1.4kb) exactly but the vector was around 4kb.
I used pET15b vector and the restriction sites are overlapping one.
Please let me know what could be the problem?

Thanks.
Aravinth.

[biocrazy]

Are you sure there is no other restriction site in the vector for the enzymes that you used?

If yes, it may undergone a non-specific digestion at some other place in the vector. You can try again by reducing the incubation time.

Good luck!!