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Efficient in silico designing of oligonucleotides for artificial gene synthesis

Garg, Abhishek D. (2009) Efficient in silico designing of oligonucleotides for artificial gene synthesis. Nature Protocols. ISSN 1750-2799

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Abstract

‘Artificial/Chemical Gene-Synthesis’ has become an important contemporary technique, which has been refined several folds owing to application of Polymerase Chain Reaction (PCR) (Engels, 2005). This procedure has been used in past for artificially synthesizing a number of genes (Engels, 2005). The principle of Artificial Gene-Synthesis has been elucidated in Figure 1 (Casimiro et al, 1997; Engels, 2005; Engels & Uhlmann, 1989; Tsuchiya et al, 2006). Technically speaking, this procedure involves two phases i.e. upstream-phase and downstream-phase. In the upstream phase, the oligonucleotides (whose sequence is derived from target gene to be synthesized) are designed manually or in silico while in the downstream phase, these designed oligonucleotides are chemically synthesized and assembled together to give stable ds-DNA duplexes/target gene. The peculiarity of these oligonucleotides is that, each forward (5’→3’) oligonucleotide overlaps with the corresponding reverse (3’→5’) oligonucleotide in around 20 terminal nucleotide-bases, thus allowing formation of an annealed-product. Current protocol focuses on the upstream-phase since it’s a success-defining phase for artificial gene-synthesis. In the current protocol, various state-of-art bioinformatics tools have been brought together to provide a comprehensive-approach towards in silico designing of oligonucleotides. Further, for the first time, DNA/mRNA secondary-structure formation analysis has been included in such a protocol.

Item Type: Article
Keywords: Oligonucleotides, DNA, in silico, Artificial Gene Synthesis
Institution: The University of Leeds
Academic Units: The University of Leeds > Faculty of Biological Sciences (Leeds)
Depositing User: Mr Abhishek Dinkarnath Garg
Date Deposited: 30 Jan 2009 09:38
Last Modified: 15 May 2014 10:57
Published Version: http://dx.doi.org/10.1038/nprot.2009.15
Status: Published
Publisher: Nature Publishing Group
Identification Number: 10.1038/nprot.2009.15
Related URLs:
URI: http://eprints.whiterose.ac.uk/id/eprint/5407

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