Restriction enzyme mapping issues

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fantastatron
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Restriction enzyme mapping issues

Post by fantastatron » Thu Jan 07, 2010 10:14 pm

Hello, I need some help determining a plasmid map...

I understand the principles and have successfully mapped RE fragments before, but I CANNOT get this one to work! I've checked that the fragment sizes are correct, and I can solve it, but only by doing things I don't think you're supposed to do!

I don't expect anyone to answer this for me (although if anyone wants to it would be VERY much appreciated), but if someone could point out where I may be going wrong...? :? I changed course after the first year and missed the basics of this, so I might be going about it in an unecessarily complicated way (although the tutorials I've looked up online don't really explain a method that differs from the one I use...)

The digests are as follows:


BamHI 4800bp
BamHI + HinDIII 3410 + 1390bp
BamHI + PvuII 2830 + 1600 + 370bp
KpnI + HinDIII 1640 + 3160bp
KpnI + PvuII 2830 + 1850 + 120bp
HinDIII + PvuII 2830 + 1760 +210bp
PvuII 2830 + 1970

I won't put all my workings out here but basically I figured that the plasmid has to contain 120, 210, 370, 1390, 1640 fragments. I figure that somehow I need to combine these to get 2830, but I can't figure out how without using multiple 120, 210 & 370 fragments, which I assume isn't allowed? (I've only seen tutorials with double digests which don't seem to do so). My best result so far is fragments of 120, 210, 370, 1390 & 2830, which adds up to 4920 :/

I've got sheets & sheets of scribbling trying to work this out & I'm literally pulling my hair out over it!

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JackBean
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Post by JackBean » Fri Jan 08, 2010 8:00 am

You can basically use some fragment twice, if you got it in two digests.
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

fantastatron
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Re: Restriction enzyme mapping issues

Post by fantastatron » Fri Jan 08, 2010 10:34 am

So does that mean in this case I can't? As 120, 210 & 370 only appear once each.

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