Bacterial artificial chromosome library for genome-wide analysis of Chinese hamster ovary cells.
Chinese hamster ovary (CHO) cell lines are widely used for scientific research and biotechnology. A CHO genomic bacterial artificial chromosome (BAC) library was constructed from a mouse dihydrofolate reductase (DHFR) gene-amplified CHO DR1000L-4N cell line for genome-wide analysis of CHO cell lines. The CHO BAC library consisted of 122,281 clones and was expected to cover the entire CHO genome five times. A CHO chromosomal map was constructed by fluorescence in situ hybridization (FISH) imaging using BAC clones as hybridization probes (BAC-FISH). Thirteen BAC-FISH marker clones were necessary to identify all the 20 individual chromosomes in a DHFR-deficient CHO DG44 cell line because of the aneuploidy of the cell line. To determine the genomic structure of the exogenous Dhfr amplicon, a 165-kb DNA region containing exogenous Dhfr was cloned from the BAC library using high-density replica (HDR) filters and Southern blot analysis. The nucleotide sequence analysis revealed a novel genomic structure in which the vector sequence containing Dhfr was sandwiched by long inverted sequences of the CHO genome.
Omasa
Takeshi
T
Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan. omasa@bio.eng.osaka-u.ac.jp
Cao
Yihua
Y
Park
Joon Young
JY
Takagi
Yasuhiro
Y
Kimura
Shuichi
S
Yano
Hidenori
H
Honda
Kohsuke
K
Asakawa
Shuichi
S
Shimizu
Nobuyoshi
N
Ohtake
Hisao
H
D000818:Animals; D016466:CHO Cells; D002874:Chromosome Mapping; D022202:Chromosomes, Artificial, Bacterial; D006224:Cricetinae; D003412:Cricetulus; D015723:Gene Library; D005822:Genetic Vectors; D016678:Genome; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D011994:Recombinant Proteins / Q000235:genetics; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics
D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase