DNA Sequencing

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magicsiew
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DNA Sequencing

Post by magicsiew » Thu Jun 10, 2010 2:14 am

To send a DNA sample from gel extraction for sequencing, I found some info that need to ligate the DNA sequence into a vector before sending. Why is it need to do this step? Can i just send directly the DNA extracted from the gel?

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JackBean
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Post by JackBean » Thu Jun 10, 2010 9:26 am

I think it may depend on type of sequenator, but if you have the classical dideoxy than you don't have to, you just need primers for your sequence. However, the beginning and the end of the sequence reaction are usually full of mistakes, so you will lack part of your sequence and that's the reason, why is it usually sequenced with some primers outside your target sequence
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

magicsiew
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Post by magicsiew » Thu Jun 10, 2010 11:51 am

So if I ligate the DNA sequence into a vector, is there any method for me to check if there is any mistake or contamination with my work? I am still quite new in practical works, and I have no real time practical on cloning yet, just worry if any contamination or error occur, might waste lots of money for the sequencing.

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JackBean
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Post by JackBean » Fri Jun 11, 2010 8:42 pm

sure, just use some primers for the vector (some vectors have sites for M13 primers, but you need to check that;)
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

maqjacob
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Re: DNA Sequencing

Post by maqjacob » Mon Jun 21, 2010 1:01 pm

I agree with JackBean

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