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- W189337169 abstract "The authors have developed a novel procedure that permits the rapid extraction and isolation of ultra-high molecular weight DNA from avian or mammalian cells using dialysis against a solution of polyethylene glycol (PEG). Cells harvested by centrifugation and washed twice in ice-cold Ca/sup + +/- and Mg/sup + +/-free phosphate buffered saline were resuspended in 5 ml 0.01 M Tris-Cl (pH 8.0); 0.001 M EDTA (TE); sodium dodecyl sulfate and proteinase K were added to final concentrations of 0.1% and 0.1 mg/ml, respectively. After incubation at 37/sup 0/C overnight, the viscous solution was transferred to a mini-collodian bag and concentrated by dialysis against 4-5 changes of 20% PEG in TE over a period of 5 hours at RT. Concentrated samples were desalted by dialysis against fresh TE for two 15 minute intervals. DNA obtained using this procedure gives A/sub 260//A/sub 280/ consistently >1.8. Analysis of DNA size using pulsed field gel electrophoresis revealed a distribution of fragments >500 Kb in length. Further measurements examined were (1) restriction enzyme digestibility, (2) ligation efficiency of restricted DNA, and (3) cloning efficiency using the lambda vector Ch21A. This novel methodology offers a valuable alternative protocol for rapid purification of ultrahigh molecular weight DNAmore » for various applications in molecular biology.« less" @default.
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- W189337169 date "1986-05-01" @default.
- W189337169 modified "2023-09-26" @default.
- W189337169 title "New and rapid procedure for the isolation of ultra-high molecular weight eukaryotic DNA" @default.
- W189337169 hasPublicationYear "1986" @default.
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