The type of DNA attachment sites recovered from nuclear matrix depends on isolation procedure used

Donev, Rossen Mintchev 2000. The type of DNA attachment sites recovered from nuclear matrix depends on isolation procedure used. Molecular and Cellular Biochemistry 214 (1) , pp. 103-110.

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Abstract

A large variety of DNA sequences have been described in nuclear matrix attachment regions. It could be most likely a result of the different methods used for their isolation. The idea about how different types of known DNA sequences (strongly attached to the nuclear matrix, weakly attached, or not attached) directly participate in anchoring DNA loops to the nuclear matrices isolated by different experimental procedures was tested in this study. Matrix-attached (M) and matrix-independent or loop (L) fractions as well as nuclear matrices were isolated using extractions of nuclei with 25 mM lithium 3,5-diiodosalicylate (LIS), 2 M NaCl, 0.65 M ammonium sulphate containing buffers followed by DNase I/RNase A digestion, or according to so designated conventional method. Using PCR-based and in vitro binding assays it was established that LIS and ammonium sulphate extractions gave similar results for the type of attachment of sequences investigated. The harsh extraction with 2 M NaCl or the conventional procedure led to some rearrangements in the attachment of DNA loops. As a result a big part of matrix attached sequences were found detached in the loop fractions. However, the in vitro binding abilities of the MARs to the nuclear matrices isolated by different methods did not change.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Medicine
Subjects:	R Medicine > R Medicine (General)
Publisher:	Springer Verlag
ISSN:	0300-8177
Last Modified:	19 Mar 2016 23:45
URI:	https://orca.cardiff.ac.uk/id/eprint/63802

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