about transformation of DNA into E.Coli

H. ROYCHOWDHURY hroychow at NMSU.EDU
Fri Aug 9 10:56:52 EST 1996


On 9 Aug 1996, Sailesh Surapureddi wrote:

> 
..snip...
> 
> 2. Initally incubate your DNA with Cells for 5min till you get  the hang
> out of your method.

How does increasing the incubation from 3 to 5 min offer any more " the
hang out of your method" 

> 
> 3. Clean your ligation mixes, since the ligation buffers usually kill your
> cells.

If ligation buffers (2uL in 50uL cells)"usually kill" the cells, why do I 
get very high degree of transformation (about 10e8 to 10e9 transformants 
per ug of plasmid) with the control ligation (un-CIP'd blunt-end vector 
w/o the insert)? This, using the standard Hanahan methods, and ligating 
one hr at RT.

..snip...




			>>>>>>>>>>>>>>>>>>>>>>>>>>>
			  Hiranya S. Roychowdhury
   			  Plant Genetic Engineering Lab.
			  Box 3GL, NM State Univ.
			  Las Cruces, NM 88003
			  Phone: (505) 646-5785
			  hroychow at nmsu.edu
			<<<<<<<<<<<<<<<<<<<<<<<<<<<




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