Detection of DNA domains in Drosophila, human, and plant chromosomes possessing mainly 50- to 150-kilobase stretches of DNA.
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.
Tchurikov
N A
NA
Department of Genome Organization, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow.
Ponomarenko
N A
NA
D000818:Animals; D002875:Chromosomes / Q000502:physiology*; D002877:Chromosomes, Human / Q000502:physiology*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D015342:DNA Probes; D004330:Drosophila / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D009693:Nucleic Acid Hybridization; D010944:Plants / Q000235:genetics*; D015183:Restriction Mapping
D015342:DNA Probes; D004247:DNA