Application of the RLGS method to large-size genomes using a restriction trapper

DNA Res. 1994;1(2):99-102. doi: 10.1093/dnares/1.2.99.

Abstract

We developed a method for producing restriction landmark genomic scanning (RLGS) profiles of large-size genomes, such as those of higher plants or amphibians using a restriction trapper. Use of the conventional RLGS method is limited to genomes smaller than 3 x 10(9) bp, because the larger genomic DNAs, especially those of more than 1 x 10(10) bp, produce high background due to incorporation of radioactivity at non-specifically damaged sites. Our new method reduces the background levels by reducing genome complexity to 1/200-1/300 using a purification step to enrich DNA fragments carrying specific restriction landmarks at their ends using a restriction trapper. This step makes it possible to obtain RLGS patterns of larger genomes. Our paper describes the practical application for the RLGS method using a restriction trapper with the pine tree genome (3 x 10(10) bp/haploid genome; Pinus koraiensis Sieb. et Zucc.) as an example.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA, Plant / chemistry
  • Electrophoresis, Gel, Two-Dimensional / methods
  • Restriction Mapping*
  • Trees / genetics*

Substances

  • DNA, Plant