Variability in the pattern of random amplified polymorphic DNA

Deepak K. Khandka, Metin Tuna, Moshe Tal, Ali Nejidat, Avi Golan-Goldhirsh

Research output: Contribution to journalArticlepeer-review

30 Scopus citations

Abstract

The random amplified polymorphic DNA (RAPD) technique is a simple method to detect DNA polymorphism. It is sensitive to reaction conditions. Small changes in the reactants' concentration cause variations in amplification products. Using DNA from Asparagus officinalis, Dactylis glomerata, Mercurialis annua and Escherichia coli, we examined variability in the amplification pattern associated with reaction constituents. An increase in the ratio of Tag DNA polymerase to DNA in the reaction increased the number of amplified fragments. Increasing the concentration of primer resulted in the amplification of low molecular weight DNA fragments, while lowering the concentration resulted in high molecular weight fragments. Subsets of amplified fragments required different concentrations of magnesium for their highest intensity. Mechanical shearing of DNA obtained by sonication led to reduction in amplification of a subset of products. Enzymatic fragmentation of DNA by restriction enzymes led to loss or gain of specific fragments, depending on the DNA, primer, and restriction enzyme. RAPD markers of pooled DNA of anonymous pedigree should be critically evaluated for frequent 'false positive' markers.

Original languageEnglish
Pages (from-to)2852-2856
Number of pages5
JournalElectrophoresis
Volume18
Issue number15
DOIs
StatePublished - 1 Dec 1997

Keywords

  • Amplification variability
  • Polymerase chain reaction
  • Random amplified polymorphic DNA
  • Reaction composition

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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