BAC end-probes

Laura F Marek lmarek at iastate.edu
Thu Aug 20 09:48:58 EST 1998


In article <Pine.OSF.3.96.980819163936.5825B-100000 at plains.NoDak.edu>, 
kalavach at PLAINS.NODAK.EDU says...
>
>Hi netters,
>i am working with BAC clones from a bean BAC library and i was wanting
>to generate end-probes. i have generated riboprobes using in vitro
>transcription. Others in the literature have generated end-probes by P
>CR
>and IPCR, and i have also tried this. most of the bands seen
>after hybridization of (what i think are) end-probes to digests of the
>parent BAC clones seem to be large (nearly 15-20 Kb). i also get multi
>ple
>bands.
>
>Does anyone have experience with this or have pointers?
>
>thanks,
>
>kal
>kalavach at plains.nodak.edu
>
When we started working with BACs a couple years ago we cloned ends for probes 
following the protocol described in the Woo et al. paper (NAR 22:4922-4931).  
More recently we have been generating end probes by direct sequencing of the 
BACs to get end sequence.  We then design primers (sometimes a step in from 
the end of the BAC is necessary to get a big enough sequence) and make probe by 
PCR.  Usually gel purify the probe and don't synthesize it hot.

Laura Fredrick Marek
Agronomy Department
Iowa State University
Ames, IA 50011

lmarek at iastate.edu




More information about the Methods mailing list