One step transformation

kang at msvax.mssm.edu kang at msvax.mssm.edu
Fri Mar 3 23:14:47 EST 1995


In article <2F55D8CF at msm.cgnet.com>, J.MAYER at CGNET.COM ("Mayer, Jorge ", 3637) writes:
>
>In article <3itgh6$g3h at server.st.usm.edu>, Glen Shearer
><gshearer at whale.st.usm.edu> writes:
>>Anyone tried the Chung et. al, PNAS.  86:2172-2175 (1989) one step
>>transformation & storage of E. coli?  As I recall, the authors use
>>a solution of PEG + MgCl (called TSS) to resuspend log phase cells
>>and get a reported 10^8 transformants per ug.
>>
>>I've tried this method with several strains and get at best 10^5
>>transformants/ug with various pUC vectors.
>>
>>I'm curious if anyone has had this work in the 10^8 per ug range and
>>if so, what is the secret technique.
>>
>>Thanks,
>>Glen
>>
>>[gshearer at whale.st.usm.edu]
>>
>
>>I used modified methods of this and usually could get 10^8 per ug.
>>I think the method was reported 1992 by Nishimura et al.
>>If you can not find the protocol send me e-mail. I have a copy in my lab.
>>One thing I want to mention is if you want to get more than 10^7 per ug DNA 
>you
>>should use DNA less than 100 pg. If DNA is to much there will be a 
>significant
>>decrease of transformation because of saturation effect.
>>Good Luck
>
>>Chulho Kang
>
>
>We have been using a method published  in NAR  22, 2857-2858 (1994) with 
>great success (2x10^9cfu/ug).  It's simplicity is incredibly attractive; in 
>principle it's just harvesting the cells at OD 0.9 instead of early-log.
>
>Jorge Mayer  <j.mayer at cgnet.com>
>
>

Hi Pal;

Sorry for mis-information. The Nishimura's paper is not 1992 but it is 1990.

I also tried 1994 NAR method without good result.

Chulho Kang



More information about the Methods mailing list