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Sizing BACs

Mon Mar 18 15:10:25 EST 1996

It would be much appreciated if those who might know could give us some
advice on determining the size of the BACs in the Texas A&M University
Arabidopsis BAC library.  We understand that they average 100 KB and 
would like to have the actual sizes of individual BACs to within 10 KB.

Is this possible using CHEF gels?  We have tried several sets of CHEF 
parameters with little success.  The DNA is smeared in the lane.  We start
with mini-prepped DNA.  Restricition-digested DNA looks fine on agarose
mini-gels performed to check the prep for degradation.  Could mini-prepped
DNA be sheared for CHEFs to work?  Is there a protocol for embedding BACs
like there is for YACs? 

The CHEF conditions we used we obtained from the WWW and were from Texas
A&M and also from Cal Tech.  

Thank you for your input.  Please reply to Zachgo at frodo.mgh.harvard.edu.

Eve Zachgo

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