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Amplification of portions of the genome in mammalian somatic cells resistant to colchicine. VI. Restriction analysis of the amplified DNA sequences

Gudkov, A.V.; Kopnin, B.P.

Genetika 19(7): 1045-1053

1983

ISSN/ISBN: 0016-6758
PMID: 6311672
Document Number: 200243
Fragments specific for the amplified regions in DNA of Djungarian hamster colchicine-resistant cells were studied after restriction endonuclease digestion. We used three different methods of detection of these fragments: a) comparison of the wild type and resistant cell DNA electroforegramms stained by ethidium bromide; b) blotting of DNA from sensitive and resistant variants onto nitrocellulose filters and their hybridization with nick-translated DNA from resistant cells, in the presence of the excess of unlabelled DNA from the wild type cells (competitive hybridization); c) investigation of autonomously replicating DNA from sensitive and colchicine-resistant sublines. The highest resolution was found using the third method. However, the competitive hybridization is evidently a more universal approach to restriction analysis of DNA amplified sequences, because it gives quite high resolution and may be used for studying both autonomously and non-autonomously replicating sequences.

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