A new method for constructing NotI linking and boundary libraries using a restriction trapper.
We have developed a novel method for constructing NotI linking and boundary libraries using a modified "solid-supported ligation primer" (restriction trapper). The restriction trapper could be used to purify the DNA fragments with a specific restriction enzyme cutting site(s) at their ends. The method uses a ligation and recutting reaction with double-stranded DNA ends of a hairpin-shaped oligolinker which is connected covalently to the surface of the latex beads. Selectivity is based on the specificity of the restriction enzyme for its recognition site, resulting in efficient purification. We applied this technique to the construction of high-quality NotI linking and NotI boundary libraries, which contain almost all the NotI sites of the genome and, in addition, are free of illegitimately ligated clones.
Hayashizaki
Y
Y
Department of Bioscience, National Cardiovascular Center Research Institute, Osaka, Japan.
Hirotsune
S
S
Hatada
I
I
Tamatsukuri
S
S
Miyamoto
C
C
Furuichi
Y
Y
Mukai
T
T
D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism*; D015723:Gene Library; D008040:Genetic Linkage; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation
D004247:DNA; D015252:Deoxyribonucleases, Type II Site-Specific; C038565:GCGGCCGC-specific type II deoxyribonucleases