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Nucleic Acids Research 2005 33(Web Server Issue):W521-W525; doi:10.1093/nar/gki380
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© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oupjournals.org

Article

Assembly PCR oligo maker: a tool for designing oligodeoxynucleotides for constructing long DNA molecules for RNA production

Roman Rydzanicz, X. Sharon Zhao and Philip E. Johnson*

Department of Chemistry, York University 4700 Keele Street, Toronto, Ontario M3J 1P3, Canada

*To whom correspondence should be addressed. Tel: +1 416 736 2100 Ext 33119; Fax: +1 416 736 5936; Email: pjohnson{at}yorku.ca

Received February 11, 2005. Revised March 8, 2005. Accepted March 8, 2005.

We describe a computer program, Assembly PCR Oligo Maker, created to automate the design of oligodeoxynucleotides for the PCR-based construction of long DNA molecules. This program is freely available at http://publish.yorku.ca/~pjohnson/AssemblyPCRoligomaker.html and has been specifically designed to aid in the construction of DNA molecules that are to be used for the production of RNA molecules by in vitro synthesis with T7 RNA polymerase. The input for Assembly PCR Oligo Maker is either the desired DNA sequence to be made or an RNA sequence. If RNA is the input, the program first determines the DNA sequence necessary to produce the desired RNA molecule. The program then determines the sequences of all the oligodeoxynucleotides necessary for a two-step assembly PCR-based synthesis of the desired DNA molecule. The oligodeoxynucleotide sequences outputted are designed to have a uniform melt temperature and are checked for regions of overlap outside of the desired priming regions necessary for the PCR reaction. The validity of the program was verified experimentally by synthesizing a 191-nt long DNA molecule using the DNA sequences suggested by the program.


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