cloning concatemers

HARDIES at THORIN.UTHSCSA.EDU HARDIES at THORIN.UTHSCSA.EDU
Mon Jan 16 15:17:15 EST 1995


To: methods-and-reagents at net.bio.net
From: sbhattac at hgmp.mrc.ac.uk (Dr. S. Bhattacharya)
Subject: cloning concatemers
Date: 15 Jan 1995 01:56:12 -0000
Sender: lpddist at mserv1.dl.ac.uk
Message-ID: <3f9vbs$25i at mserv1.dl.ac.uk>
Original-To: methods at dl.ac.uk

Shoumo Bhattacharya wrote:

> Can anyone guide me to a reference describing how to clone
> concatemers of an operator site without getting inversions,
> i.e. all units of the concatemer in the same direction.

In most strains, the inverted repeats are selected against because the
replication apparatus can't handle them.  So if you just ligate
monomers at 10:1 molar ratio or so with vector and pick the larger
inserts, you'll get 2x and 3x head to tail insertions.  It's hard
to get longer concatemers because the statistics of getting all
fragments the same way by chance is against you.  If you need
a long concatemer, I'd consider synthesizing overlapping oligos
with ends that enforce head to tail joining during ligation.

Steve Hardies, Dept. of Biochemistry, Univ. of Texas HSC at San
Antonio
Hardies at uthscsa.edu




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