Help! Plz. Cloning

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Help! Plz. Cloning

Post by sem.meln » Wed May 03, 2006 8:49 am

I cloned a 250 bp long vector into 5 kb plasmid. When I checked the insert sequence it turned out there is a mutated fragment of 10 bp length. I cut out the vector to clone a new one, but when I run the cut plasmid on gel- I can not distinguish where the cut and where not cut plasmid! (250 bp is too short to see the difference). Of course, I can cut the entire bend, try to ligate a new insert and hope ligation occurs, but how I will check whether mutation disappeared?
Is the some way to correct the mutation?
Thank you in advance!

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Post by LilKim » Thu May 04, 2006 2:57 pm


this sounds complicated!

Gosh, I'd just start over (if thats an option)!!!!

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Post by canalon » Thu May 04, 2006 5:19 pm

You clone a vector in a plasmid?

Usually the plasmid is called the vector, and whatever you clone is the insert...

But to answer your question: I woud rateher start over if possible, since correcting is usually more risky than anything else. What can of insert do you have? because that could explain were your problem is. Because it is unlikely that a simple ligation can have caused mutations.

Science has proof without any certainty. Creationists have certainty without
any proof. (Ashley Montague)

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