Restriction site - (Jan/09/2007 )
I need to realise a restriction on a plasmid with MscI. There is 2 restriction site on this plasmid but I am interested just by one them.
Is someone know how can I do my restriction to have a single restriction and not two.
Can I decrease the time of restriction to have partial digestion and y chance obtain the good one???
Or do you have any other idea to do that?
Thanks a lot
Have a nice day
u may decrease the reaction time. i would try 20 minutes.
I would decrease time, (20 minutes) purify the partial digested, cut with the second enzyme and pray that the digested product has the right size.
If you are unlucky, the site that you don't want to digest will be mor easily digested, and you will have to perform a site directed mutagenesis..
But maybe you are lucky and at least have 50% of the right digestion, so try first the partial digestion.
Hello thank you for your interest,
I Try different time of reaction and now I just put on gel and wait the result. I cross my finger!!!