Plasmid loss after restreaking

Graham Dellaire popa0206 at PO-Box.McGill.CA
Tue Jul 18 22:14:08 EST 1995


>   ddh at unlinfo.unl.edu (dinakar bhattramakki) writes:
>  Hi netters,
>  
>  I have a problem which I think was discussed in detail very recently. 
>  Could any one post the conclusion of that discussion?
>  "PROVERB IS NOT A PROVERB UNLESS YOU EXPERIENCE IT"
>  
>  My problem: I transformed Sure18 cells with recombinant pBluescript 
>  plasmid. Selected the white transformants on LB+Amp+X-gal+IPTG plates.
>  I did a master plate of half a dozen transformants. I was unable to 


Didi you restreak from the master plate...?

Often what will happen is that when you just pick a mixed colony (from your master plate...
unless you streaked your first pick and then used a single ampR colony for your master plate)
that when grown in broth will end up with a large population of bacteria without plasmid and they 
willl out grow your plasmid carrying bacteria.  End result your plasmid is lost from culture


Graham

So bottom line always restreak your first pick and then make a master plate from the restreaked 
"pure" colonies.


Graham Dellaire

                                    Red Cross, Research		
McGill University                   Montreal Blood Services	  	
Faculty of Medicine                 3131 Sherbrooke St. East         
Div. of Experimental Medicine       Montreal, QC, Canada           
E-mail: popa0206 at po-box.mcgill.ca   H1W 1B2			   
B2XE at musicb.mcgill.ca							   
WWW Page: http://www.medcor.mcgill.ca/EXPMED/expmed.html	
Fax: (514) 525 0881							   	
Voice: (514) 527 1501 ext 175 						  
_______________________________________________________________________





More information about the Methods mailing list