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Nucleic Acids Research Advance Access published online on October 11, 2007
Nucleic Acids Research, doi:10.1093/nar/gkm857
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© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Methods Online

In vivo construction of recombinant molecules within the Caenorhabditis elegans germ line using short regions of terminal homology

Benedict J. Kemp, Julia Hatzold, Laura A. Sternick, Joshua Cornman-Homonoff, Jessica M. Whitaker, Pamela J. Tieu and Eric J. Lambie*

Department of Biological Sciences, Dartmouth College, Hanover, NH 03755, USA

* To whom correspondence should be addressed. Tel: +1 603 646 3875; Fax: +1 603 646 1347; Email: eric.j.lambie{at}dartmouth.edu

Received August 30, 2007. Revised September 25, 2007. Accepted September 26, 2007.

Homologous recombination provides a means for the in vivo construction of recombinant DNA molecules that may be problematic to assemble in vitro. We have investigated the efficiency of recombination within the Caenorhabditis elegans germ line as a function of the length of homology between recombining molecules. Our findings indicate that recombination can occur between molecules that share only 10 bp of terminal homology, and that 25 bp is sufficient to mediate relatively high levels of recombination. Recombination occurs with lower efficiency when the location of the homologous segment is subterminal or internal. As in yeast, recombination can also be mediated by either single- or double-stranded bridging oligonucleotides. We find that ligation between cohesive ends is highly efficient and does not require that the ends be phosphorylated; furthermore, precise intermolecular ligation between injected molecules that have blunt ends can also occur within the germ line.

The authors wish it to be known that, in their option, the third and fourth and fourth authors should be regarded as join First Authors.


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