Rapid cloning of multiple amplified nucleotide sequences from human neuroblastoma cell lines by phenol emulsion competitive DNA reassociation.
A protocol for the rapid cloning of many DNA fragments from an amplified genomic region is described. The procedure is based on a modification of the phenol-emulsion reassociation technique (PERT) previously used to clone DNA fragments missing from a chromosomal deletion [Kunkel et al., Proc. Natl. Acad. Sci. USA 82 (1985) 4778-4782]. The procedure was used to construct recombinant libraries in the plasmid pBR322 which were highly enriched for amplified sequences from two neuroblastoma cell lines, CHP-126 and IMR-32. Many new amplified DNA fragments were isolated from these libraries, indicating that the PERT methodology should be of general use in isolating amplified DNA from other cell lines and tumors.
D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods*; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification*; D004274:DNA, Recombinant / Q000302:isolation & purification*; D004655:Emulsions; D006801:Humans; D009447:Neuroblastoma / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D019800:Phenol; D010636:Phenols
D004273:DNA, Neoplasm; D004274:DNA, Recombinant; D004655:Emulsions; D010636:Phenols; D019800:Phenol