Large-scale, cost-effective screening of PCR products in marker-assisted selection applications

Weikuan Gu, N. F. Weeden, J. Yu, D. H. Wallace

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

A simple, PCR-based method has been developed for the rapid genotyping of large numbers of samples. The method involves a alkaline extraction of DNA from plant tissue using a slight modification of the procedure of Wang et al. (Nucleic Acids Res 21:4153-4154, 1993). Template DNA is amplified using allelespecific associated primers (ASAPs) which, at stringent annealing temperatures, generate only a single DNA fragment and only in those individuals possessing the appropriate allele. This approach eliminates the need to separate amplified DNA fragments by electrophoresis. Instead, samples processing the appropriate allele are identified by direct staining of DNA with ethidium bromide. Total technician time required for extraction, amplification and detection of 96 samples is about 4 h, and this time requirement can be reduced by automation. Excluding labor, cost per sample is less than $0.40. The method is tested using the codominant isozyme marker, alcohol dehydrogenase (Adh-1) gene in pea (Pisum sativum), and applied to the screening of photoperiod genes in common bean (Phaseolus vulgaris L.).

Original languageEnglish (US)
Pages (from-to)465-470
Number of pages6
JournalTheoretical And Applied Genetics
Volume91
Issue number3
DOIs
StatePublished - Aug 1 1995
Externally publishedYes

Fingerprint

marker-assisted selection
screening
Costs and Cost Analysis
Polymerase Chain Reaction
Peas
DNA
Alleles
Plant DNA
Phaseolus
Ethidium
Alcohol Dehydrogenase
Automation
Photoperiod
alleles
sampling
Nucleic Acids
annealing
Isoenzymes
Genes
technicians

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Agronomy and Crop Science
  • Genetics

Cite this

Large-scale, cost-effective screening of PCR products in marker-assisted selection applications. / Gu, Weikuan; Weeden, N. F.; Yu, J.; Wallace, D. H.

In: Theoretical And Applied Genetics, Vol. 91, No. 3, 01.08.1995, p. 465-470.

Research output: Contribution to journalArticle

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