Application of the RLGS method to large-size genomes using a restriction trapper.
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.
Okuizumi
H
H
Gene Bank, RIKEN Tsukuba Life Science Center, Ibaraki, Japan.
Okazaki
Y
Y
Sasaki
N
N
Muramatsu
M
M
Nakashima
K
K
Fan
K
K
Tano
H
H
Ohba
K
K
Hayashizaki
Y
Y
D018744:DNA, Plant / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D015183:Restriction Mapping; D014197:Trees / Q000235:genetics*
D018744:DNA, Plant