Preferential insert orientation

Paul N Hengen pnh at fcs260c2.ncifcrf.gov
Wed Dec 29 15:39:27 EST 1993


In article <gregoryh-271293155836 at 128.249.14.251>
gregoryh at bcm.tmc.edu (Gregory R. Harriman) writes:

> What can cause inserts with the same restriction site on both ends to
> ligate in only one direction? Any advice or helpful suggestions would be
> appreciated!

1. Secondary structure of the DNA fragments may prohibit the ligation in one
particular orientation, ie., one end of each molecule is much more frequently
ligated first.

2. The transformants of both types of molecules are never seen because the
insert may be expressed from a vector promoter in only one orientation and the
product(s) are lethal to the cell.

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* Paul N. Hengen, Ph.D.                           /--------------------------/*
* National Cancer Institute                       |Internet: pnh at ncifcrf.gov |*
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