How to modify a DNA construct?

Gary K. gtk10583 at glaxo.com
Mon Feb 12 07:44:50 EST 1996


> af930 at FreeNet.Carleton.CA (Changcun Jia) wrote:
> 
> >I would like to insert a 30 bp nucleotide into a construct. Anybody has
> >any suggestion on how to do this?

In article <311d7cdb.42062718 at news.std.com>, rrohan at world.std.com (Richard
Rohan) wrote:

> Depends on if there is a unique restriction site at this location.  If
> so, then you could use and adapter oligonucleotide.  Otherwise you
> could PCR with oligos containing the 30 bp that you want to insert.
> 
> Rich Rohan
>

The PCR route is the fastest but still requires a suitable restriction
site for subcloning back into the original construct for replacement. 
Another option is to make ssDNA of your target plasmid, add your primer
with the 30 bp insert and synthesize your new strand.  There  are kits
available for making this easy.

Gary



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