Open AccessDetection of DNA domains in Drosophila, human, and plant chromosomes possessing mainly 50- to 150-kilobase stretches of DNA.
Author(s) -
N. A. Tchurikov,
Н. А. Пономаренко
Publication year - 1992
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.89.15.6751
Subject(s) - dna , microbiology and biotechnology , biology , gel electrophoresis , southern blot , agarose gel electrophoresis , nucleic acid thermodynamics , in vitro recombination , dna sequencing , restriction fragment , circular bacterial chromosome , molecular cloning , genetics , gene , base pair , complementary dna , base sequence
We have used pulsed-field gel electrophoresis of undigested DNA prepared by cell lysis in agarose with proteinase K detergent treatment and found a resolvable DNA fraction, denoted forum DNA (f-DNA). By changing the pulsed-field gel pulse length from 25 to 4500 sec, to obtain optimal separation in different ranges, we have found f-DNA to occupy a rather broad zone from 2 megabases to 10 kilobases (kb), but mainly at a range between 50 and 150 kb. f-DNA seems to appear as a result of nonrandom spontaneous degradation during cell treatment. The terminal regions of f-DNA segments have been cloned by using a jumping library. The molecular analysis of unique DNA sequence from an anonymous Drosophila DNA segment led to the conclusion that f-DNA appears as a result of nonrandom chromosomal DNA cleavage within sensitive regions that occupy a few kilobases. This conclusion was confirmed by detection of rather discrete hybridization bands on pulsed-field gel Southern blots in a region of good separation of undigested f-DNA after hybridization with different unique and repetitive probes. We propose that f-DNA segments may correspond to some regular higher-order structures in the eukaryotic chromosomes.