Generation of high-density DNA markers from yeast artificial chromosome DNA by single unique primer-polymerase chain reaction.
We have developed a method for the whole sequence amplification of yeast artificial chromosome (YAC) DNA excised from preparative pulsed-field gel electrophoresis using single unique primer-polymerase chain reaction procedures. We used seven contiguous YAC clones, which span 2 Mbp of the Huntington disease gene region on 4p16.3, to amplify the YAC DNAs. The average size of the amplified DNA was approximately 300 bp long, and 12 DNA markers located on the YAC clones positively hybridized with these amplified products, implying that the sequences of the YAC clones were comprehensively amplified by our procedures. These amplified YAC DNAs greatly facilitate the characterization of YAC clones, leading to the detailed analysis of the defined chromosomal region.
Hadano
S
S
Genome Analysis Laboratory, Imperial Cancer Research Fund, London, England.
Ishida
Y
Y
Bates
G P
GP
Nagayama
T
T
Kanazawa
I
I
Lehrach
H
H
Ikeda
J
J
D001483:Base Sequence; D015139:Blotting, Southern; D018244:Chromosomes, Artificial, Yeast; D002894:Chromosomes, Human, Pair 4; D017931:DNA Primers; D016521:Electrophoresis, Gel, Pulsed-Field; D005819:Genetic Markers / Q000235:genetics*; D006801:Humans; D006816:Huntington Disease / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers; D005819:Genetic Markers