[35] Strategies for mapping and cloning macroregions of mammalian genomes

Smith, Cassandra L.; Lawrance, Simon K.; Gillespie, Gerald A J; Cantor, Charles R.; Weissman, Sherman M.; Collins, Francis S.

doi:10.1016/s0076-6879(87)51038-2

Cited by 98 publications

(52 citation statements)

References 19 publications

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“…High-Molecular-Weight DNA Preparation and Linearization of Circular Extrachromosomal DNAs High-molecular-weight DNA was prepared as described by Smith et al (1987). Using an agarose plug mold (Bio-Rad, Richmond, CA), -1.5 X 107 cells/ml in phosphate-buffered saline were embedded in molten agarose (InCert agarose, FMC, Rockland, ME).…”

Section: Cytogenetic Analysismentioning

confidence: 99%

Double minute chromosomes carrying the human multidrug resistance 1 and 2 genes are generated from the dimerization of submicroscopic circular DNAs in colchicine-selected KB carcinoma cells.

Schoenlein

Shen

Barrett

et al. 1992

MBoC

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This study characterizes amplified structures carrying the human multidrug resistance (MDR) genes in colchicine-selected multidrug resistant KB cell lines and strongly supports a model of gene amplification in which small circular extrachromosomal DNA elements generated from contiguous chromosomal DNA regions multimerize to form cytologically detectable double minute chromosomes (DMs). The human MDR1 gene encodes the 170-kDa Pglycoprotein, which is a plasma membrane pump for many structurally unrelated chemotherapeutic drugs. MDR1 and its homolog, MDR2, undergo amplification when KB cells are subjected to stepwise selection in increasing concentrations of colchicine. The structure of the amplification unit at each step of drug selection was characterized using both highvoltage gel electrophoresis and pulsed-field gel electrophoresis (PFGE) techniques. An 890-kb submicroscopic extrachromosomal circular DNA element carrying the MDR1 and MDR2 genes was detected in cell line KB-ChR-8-5-1 1, the earliest step in drug selection in which conventional Southern/hybridization analyses detected MDR gene amplification. When KB-ChR-8-5-11 was subjected to stepwise increases in colchicine, this circular DNA element dimerized as detected by PFGE with and without digestion with Not 1, which linearizes the 890-kb amplicon. This dimerization process, which also occurred at the next step of colchicine selection, resulted in the formation of cytologically detectable DMs revealed by analysis of Giemsa-stained metaphase spreads.

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Section: Cytogenetic Analysismentioning

confidence: 99%

Double minute chromosomes carrying the human multidrug resistance 1 and 2 genes are generated from the dimerization of submicroscopic circular DNAs in colchicine-selected KB carcinoma cells.

Schoenlein

Shen

Barrett

et al. 1992

MBoC

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“…Hybridization was carried out at 65°C for 1 Assessment of recovery and purity of the isolated fragment Two methods were employed to estimate the recovery and purity of the isolated fragment. One was to amplify the flow-through, wash fractions and the ribonuclease eluate from the avidin matrix using primers specific for ,B-globin genes and other randomly chosen genes.…”

Section: Methodsmentioning

confidence: 99%

“…The availability of linking libraries (1,2) and the feasibility of separating large size fragments obtained by partial digestion (3,4,5,6) have made large scale restriction maps an attainable objective. Cosmid (7) and yeast artificial chromosome (YAC) (8) cloning techniques are useful for ordering successively larger pieces of the genome.…”

Section: Introductionmentioning

confidence: 99%

Selective enrichment of a large size genomic DNA fragment by affinity capture: an approach for genome mapping

1990

Trends in Genetics

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A method to enrich large size DNA fragments obtained by digestion with rare cutting restriction endonucleases was developed and applied for the isolation of a 150 kb Sfil fragment containing the ,B-globin gene cluster.The digested DNA is rendered single stranded at the ends by diffusing a strand specific exonuclease into an agarose plug containing DNA. The plug is melted and solution hybridization is then performed with a bridge RNA containing specific sequences from the end of a desired fragment linked to a common probe sequence. The common probe sequence is annealed to a biotinylated RNA and the resulting tripartite hybrid is retained onto a solid matrix containing avidin and specifically released by ribonuclease action.

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“…Assuming that CpG islands in chromosome have average length of 1,000 bp, make up 1% of total DNA, have an average base composition of 65% G+C compared to 40% of bulk DNA and show no deficiency of CpG in the island region, Lindsay and Bird (1987) predicted that 74% of sites for CpG enzymes recognizing 6 bases (such as SaclI, BssHII and EagI) and 89 % of sites for those recognizing 8 bases (such as NotI)should be located in CpG islands. This estimation have led us to the development of strategies that allow specific cloning of potential CpG islands and hence of genes using the sites of CpG enzymes as indicator (Smith et al, 1987;Frischauf, 1989;Kusuda et al, 1989).…”

Section: Introductionmentioning

confidence: 99%

Over 80% ofNotI sites are associated with CpG rich islands in the sequenced human DNA

et al. 1990

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SummaryClones containing DNA segments linked to NotI sites are not only useful for ordering the NotI fragments fractionated by pulsed field gel, but also valuable in the search of unknown genes, because they often contain the CpG rich islands and genes related with them, To know the probability of association of NotI sites with CpG rich islands, we screened 5,188 sequences accumulated in DNA data base for the presence of NotI site and examined the distribution of CpGs around them. The sequential calculation of G+C content and frequency of CpG occurrence at each nucleotide position identified the CpG rich domains close to NotI sites in 77 sequences, which corresponds to 84% of total number of candidate sequences. This frequency is consistent well with the prediction that 89 % of NotI sites in mammalian genome are likely to be present in CpG rich islands and would stress the importance of cloning of NotI linking sequences for direct isolation of desired genes. Furthermore, 63 islands newly identified in this study should provide a clue for understanding the transcriptional regulation of associated genes.

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[35] Strategies for mapping and cloning macroregions of mammalian genomes

Cited by 98 publications

References 19 publications

Double minute chromosomes carrying the human multidrug resistance 1 and 2 genes are generated from the dimerization of submicroscopic circular DNAs in colchicine-selected KB carcinoma cells.

Double minute chromosomes carrying the human multidrug resistance 1 and 2 genes are generated from the dimerization of submicroscopic circular DNAs in colchicine-selected KB carcinoma cells.

Selective enrichment of a large size genomic DNA fragment by affinity capture: an approach for genome mapping

Over 80% ofNotI sites are associated with CpG rich islands in the sequenced human DNA

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