pCaSpeR-hs cloning problem

Duc Nguyen defrankenstein at yahoo.com
Fri Apr 11 08:35:54 EST 2003


Chris,

Sounds like you have a contamination with pBluescript. Somewhere
somehow, I get that often. I usually do PCR colony screen to get the
right construct, using a P vector-specific primer together with one in
my insert.

Chris Jones <6kuritaro at earthlink.net> wrote in message news:<6kuritaro-CD5F10.20302503042003 at nnrp02.earthlink.net>...
> Hi there --
> 
> I'm hoping someone has an explanation (or even better, a solution) based 
> on experience:
> 
> My student is trying to subclone a 4.1 kb Xba fragment into pCaSpeR-hs 
> (ca. 8.8 kb) in preparation for egg injections. The problem is that when 
> we ligate and transform bacteria, the resulting plasmids are either just 
> recircularized vector (no surprise there) or uniformly too small -- each 
> of these consists of a 4.1 kb fragment and a (roughly) 2.7 kb fragment 
> when re-cut with Xba. What's happened to the pCaSpeR-hs?
> 
> The insert has already been subcloned into another vector, so it seems 
> to be tractable. Has anyone ever run across something like this? We've 
> tried this several times, always with the same results. As you can 
> imagine, it's very frustrating (especially for my student).
> 
> Thanks for any advice.




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