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Restriction Digest Question


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2 replies to this topic

#1 PandaCreamPuff

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Posted 04 July 2010 - 01:21 AM

Hi all

I have two sequences of about 700bp, and I would like to see if a particular RFLP enzyme can differentiate them from each other on a gel. Is there a programme that I can use to tell me which enzyme I should use? I am currently tackling NEBcutter but not having much luck.

Thanks all!

#2 Curtis

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Posted 04 July 2010 - 02:52 AM

you need a 'Restriction Site Mapper'

http://www.restrictionmapper.org/

#3 Trof

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Posted 07 July 2010 - 07:00 AM

First you should align these two sequences and see where they differ. Then you paste them separately to NEBcutter and find the position of the differencies, zoom in, so you can see enzymes recognising your region. Compare between the two sequences if there is an enzyme, that cuts only one of them. Then make a custom digest with that enzyme and choose "View gel" for each of the sequences. You can say if it's possible to distinguish them on the gel.

I did search for enzyme distinguishing my wild-type and mutant gene like this.

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