Karnik, A., Karnik, R. and Grefen, C. (2013) SDM-Assist software to design site-directed mutagenesis primers introducing “silent” restriction sites. BMC Bioinformatics, 14, 105. (doi: 10.1186/1471-2105-14-105) (PMID:23522286) (PMCID:PMC3644487)
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Abstract
Background: Over the past decades site-directed mutagenesis (SDM) has become an indispensable tool for biological structure-function studies. In principle, SDM uses modified primer pairs in a PCR reaction to introduce a mutation in a cDNA insert. DpnI digestion of the reaction mixture is used to eliminate template copies before amplification in E. coli; however, this process is inefficient resulting in un-mutated clones which can only be distinguished from mutant clones by sequencing. Results: We have developed a program – ‘SDM-Assist’ which creates SDM primers adding a specific identifier: through additional silent mutations a restriction site is included or a previous one removed which allows for highly efficient identification of ‘mutated clones’ by a simple restriction digest. Conclusions: The direct identification of SDM clones will save time and money for researchers. SDM-Assist also scores the primers based on factors such as Tm, GC content and secondary structure allowing for simplified selection of optimal primer pairs.
| Item Type: | Articles |
|---|---|
| Status: | Published |
| Refereed: | Yes |
| Glasgow Author(s) Enlighten ID: | Grefen, Dr Christopher and Karnik, Dr Rucha |
| Authors: | Karnik, A., Karnik, R., and Grefen, C. |
| College/School: | College of Medical Veterinary and Life Sciences > School of Molecular Biosciences |
| Journal Name: | BMC Bioinformatics |
| Publisher: | BioMed Central |
| ISSN: | 1471-2105 |
| ISSN (Online): | 1471-2105 |
| Copyright Holders: | Copyright © 2013 Karnik et al. |
| First Published: | FIrst published in BMC Bioinformatics 14: 105 |
| Publisher Policy: | Reproduced under a Creative Commons License |
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