Hello everybody!

I would need some help to figure out how to separate the insert of a BAC clone from the backbone vector. I have tried the following procedure but I could only recover a few ng of insert (which is not enough for downstream applications):

1/ cut with restrictions enzymes between the insert and the backbone

2/ run the resulting restriction reaction on PFGE

3/ cut out the band corresponding to the cut insert

4/ purify DNA from gel thanks to agarase (the insert is ~200kb long so I can not purify it on a commercial column)

Could anyone suggest an alternative protocol?

Thank you in advance!

Anne-Sophie