Strategies to Improve Efficiency and Specificity of Degenerate Primers in PCR View Full Text


Ontology type: schema:Chapter     


Chapter Info

DATE

2017

AUTHORS

Maria Jorge Campos , Alberto Quesada

ABSTRACT

PCR with degenerate primers can be used to identify the coding sequence of an unknown protein or to detect a genetic variant within a gene family. These primers, which are complex mixtures of slightly different oligonucleotide sequences, can be optimized to increase the efficiency and/or specificity of PCR in the amplification of a sequence of interest by the introduction of mismatches with the target sequence and balancing their position toward the primers 5'- or 3'-ends. In this work, we explain in detail examples of rational design of primers in two different applications, including the use of specific determinants at the 3'-end, to: (1) improve PCR efficiency with coding sequences for members of a protein family by fully degeneration at a core box of conserved genetic information, with the reduction of degeneration at the 5'-end, and (2) optimize specificity of allelic discrimination of closely related orthologous by 5'-end degenerate primers. More... »

PAGES

75-85

References to SciGraph publications

Identifiers

URI

http://scigraph.springernature.com/pub.10.1007/978-1-4939-7060-5_4

DOI

http://dx.doi.org/10.1007/978-1-4939-7060-5_4

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1085581242

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/28540700


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