A new band after CIPing

Antonin Tutter atutter at aim.salk.edu
Sun Oct 4 00:18:19 EST 1998

>I cut my vector (pIVET8) with BglII, gel purify it (Nusieve Agarose), CIP
>it. To deactivate the CIP, I used Proteinake K an d SDS and incubated at
>55 C for 30min. Phenol-chloroforme extraction then precipitation with NaAC
>and ethanol.
>My surprise is the apparition of a new band with a high molecular weight
>(>30KB) while my plasmid is around 10 Kb. I don't know what happened. did
>my digested plasmid polymerize, I really have no clue.
>So I was wondering if anyone had that sort of weird size enhancement or
>not? and is there any explanation to that phenomenon?

Why don't you simply heat kill the BglII, CIP treat and THEN gel purify?  As
for the high mw band, perhaps one of your reagents is contaminated with DNA.

Antonin Tutter
Salk Institute for Biological Studies
10010 N. Torrey Pines Rd.
La Jolla, CA  92037
email:  atutter at aim.salk.edu
web:  http://www-biology.ucsd.edu/~atutter/

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