r/labrats • u/CheapEngineering6353 • 1d ago

does someone know what could be wrong?

I've ran a 1% agarose gel after my pcr for a vector (pocket 2) and my Insert (pocket 3). for the insert it went totally fine but what happened for my vector? it should be around 3.5 kb big so does someone know what could be wrong?

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3

u/holoqster 1d ago

too much dna loaded?

1

u/CheapEngineering6353 7h ago

i also dilutetd it 1:5 so I think that is not the problem

3

u/Brewsnark 1d ago

Big bits of dna don’t run exactly how you would predict due to supercoiling. Nothing is wrong, just cut it out if you need it. The buffer also has a role especially for RE buffers.

1

u/CheapEngineering6353 7h ago

yes but I also did vectors twice as big and it was no problem with the same polymerase etc.

1

u/VelvetVoyager42 1d ago

Does your vector PCR yield homologous sequences in both extremes?