DNA sequencing confusion

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stopherlogic
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DNA sequencing confusion

Post by stopherlogic » Mon Jan 05, 2009 9:58 pm

I was wondering when DNA is sequenced eg in the human genome project BACs or other vectors are used. Every time a piece of DNA is sequenced do you have to sequence the vector also? And if you do isn't this a waste of time?

blcr11
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Re: DNA sequencing confusion

Post by blcr11 » Mon Jan 05, 2009 11:53 pm

You don't bother to re-sequence the vector. It isn't necessary.

stopherlogic
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Post by stopherlogic » Tue Jan 06, 2009 12:24 pm

Thank you, does this mean you must separate your target DNA from the vector before sequencing using a restriction enzyme?

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Re: DNA sequencing confusion

Post by blcr11 » Tue Jan 06, 2009 2:35 pm

No. You select a set of sequencing primers to assure yourself that you get all of the insert material. Unless you deliberately use vector sequence as a primer (which you might do to to confirm the boundaries of your insert), you won't be picking up vector sequence except as a kind of run-off from a primer designed against insert which happens to lie near one or the other boundary. When you start seeing vector sequence, you stop reading sequence any further in that direction. Once you've hit vector in both directions, and you've overlapped everything else in between, you know you're finished. Most likely, you will only be interested in keeping your insert-sequence and will edit out any vector sequence you got in the process.

stopherlogic
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Re: DNA sequencing confusion

Post by stopherlogic » Tue Jan 06, 2009 6:22 pm

That has answered everything I need to know, thank you.

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