help in identefying a sequence within a sequence

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shachtal
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help in identefying a sequence within a sequence

Post by shachtal » Mon Aug 06, 2007 4:37 pm

i got a plasmid from a friend of mine
it is supposed to have a gfp protein under cmv promotor

the thing is
i lost contact with the dude that gave me this plasmid
but i do know for a fact that this plasmid contains GFP under some eokarayotic promotor, cause whenever i transfect it into culture cells. they start to shine like the green man from chernobil.

my problem is, i want to create a nice graphical map with all the restriction sites of the plasmid, and all of the functional elements on it (such as oris, GFP, and so on and so on.....)
i know how to create a restriction site map.... but...
how do i find those functional sequences(such as gfp) inside my plasmid?
i tried looking for parts of gfp sequence inside my plasmid, but a simple "find search" in word won't work

anyway, i hope you will help me, because i do believe there is a simple solution for this problem :)

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canalon
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Post by canalon » Tue Aug 07, 2007 2:31 am

So you just have a text file of the sequence. what yo need is just a software that will be able to read your sequence and will put all the restriction site at the right place.

In I think the New England Biolabs website can do that for you for most of the known restriction enzyme. And Blst (on NCBi for example) can easily locate the GFP in your sequence.

Of course molecular biology suite of software can do both easily (Vector NTI is free, lasergene and many other are also available).
Patrick

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

blcr11
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Post by blcr11 » Tue Aug 07, 2007 1:21 pm

I'm assuming you have the sequence of the vector + GFP as well as just the GFP sequence. You can enter them into the SIM tool of the Expasy proteomics server ( http://www.expasy.ch/tools/sim-nucl.html ) and align the two sequences. You should end up aligning GFP with GFP within the vector, which is what you wanted, if I understand you correctly.

If you want "pretty" restriction maps, things like VectorNTI work (it's only free for academics, though). I think the NEB webtools have already been mentioned, but I don't think that will give you circular maps, though I could be wrong about that. Still worth checking it out. The Expasy server also has a mapping tool, though again, it will only give you a linear map, I think.

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