A tandem insertion vector for large-scale preparation of nucleosomal DNA.
We developed a novel nucleosome DNA template vector, pWMD01, which is optimized for the large-scale preparation of nucleosomal DNA. By using restricted digestion by SapI or EarI within its multicloning site, multiple half-nucleosome DNA units can be introduced unidirectionally into the vector at each subcloning step. Through this method, we constructed a plasmid that has 18 tandem repeats of a half-nucleosome 90-bp DNA unit containing c-Myb-binding sites in two subcloning cycles. This method enables the rapid, large-scale preparation of nucleosomal DNA with crystallization-grade quality.
RIKEN Systems and Structural Biology Center, Tsurumi, Yokohama 230-0045, Japan.
Masatoshi
M
Wakamori
Takashi
T
Umehara
Shigeyuki
S
Yokoyama
D001665:Binding Sites; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D005821:Genetic Techniques; D005822:Genetic Vectors / Q000096:biosynthesis*; D016254:Mutagenesis, Insertional; D009707:Nucleosomes / Q000235:genetics*; D020598:Proto-Oncogene Proteins c-myb / Q000378:metabolism; D020080:Tandem Repeat Sequences
D009707:Nucleosomes; D020598:Proto-Oncogene Proteins c-myb; D004247:DNA