Isolation of High-molecular-weight DNA from Mammalian Cells Using Formamide

2006 ◽  
Vol 2006 (1) ◽  
pp. pdb.prot3225 ◽  
Author(s):  
Joseph Sambrook ◽  
David W. Russell
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Mammalian Cells ◽  
High Molecular Weight Dna

scholarly journals Isolation of High-Molecular-Weight DNA from Mammalian Cells

1973 ◽  
Vol 36 (1) ◽  
pp. 32-38 ◽  
Author(s):  
Maria Gross-Bellard ◽  
Pierre Oudet ◽  
Pierre Chambon
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Mammalian Cells ◽  
High Molecular Weight Dna

Isolation of high molecular weight DNA from wholeEuglena cells

PROTOPLASMA ◽  
1987 ◽  
Vol 141 (2-3) ◽  
pp. 139-148 ◽  
Author(s):  
Christine Mederic ◽  
Odile Bertaux ◽  
J. D. Rouzeau ◽  
R. Valencia
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
High Molecular Weight Dna

A general method of isolating high molecular weight DNA from methanogenic archaea (archaebacteria)

1992 ◽  
Vol 38 (1) ◽  
pp. 65-68 ◽  
Author(s):  
Ken F. Jarrell ◽  
David Faguy ◽  
Anne M. Hebert ◽  
Martin L. Kalmokoff
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Thermal Denaturation ◽  
Dna Isolation ◽  
Methanogenic Archaea ◽  
Restriction Enzymes ◽  
Mole Percent ◽  
High Molecular Weight Dna ◽  
Isolation Methods ◽  
General Method

High molecular weight DNA was readily isolated from all methanogens treated, as well as from thermophilic anaerobic eubacteria, by grinding cells frozen in liquid N2, prior to lysis with SDS. DNA can subsequently be purified by the usual phenol–chloroform extractions. The procedure yields DNA readily cut by restriction enzymes and suitable for oligonucleotide probing, as well as for mole percent G + C content determination by thermal denaturation. The method routinely yields DNA of high molecular weight and is an improvement over DNA isolation methods for many methanogens, which often involve an initial breakage of the cells in a French pressure cell. Key words: methanogens, archaebacteria, archaea, DNA isolation.

Role of proteolysis in apoptosis: involvement of serine proteases in internucleosomal DNA fragmentation in immature thymocytes

1993 ◽  
Vol 71 (9-10) ◽  
pp. 488-500 ◽  
Author(s):  
Valerie M. Weaver ◽  
Boleslaw Lach ◽  
P. Roy Walker ◽  
Marianna Sikorska
Keyword(s):  
Molecular Weight ◽  
Serine Protease ◽  
High Molecular Weight ◽  
Protease Inhibitors ◽  
Dna Fragmentation ◽  
Dna Cleavage ◽  
Apoptotic Cell ◽  
Dependent Manner ◽  
Serine Protease Inhibitors ◽  
High Molecular Weight Dna

Three chemically distinct serine, but not cysteine, protease inhibitors (phenylmethylsulphonyl fluoride, N-tosyl-L-phenylalanylchloromethyl ketone and 3,4-dichloroisocoumarin) prevented, in a dose-dependent manner, the characteristic apoptotic internucleosomal DNA cleavage (DNA ladder) typically observed in thymocytes in response to dexamethasone and teniposide VM-26. This effect was not the result of a direct inhibition of the Ca2+, Mg2+-dependent endonuclease, since oligonucleosomal DNA cleavage occurred in the presence of these inhibitors in isolated nuclei. The proteolytic step occurred at a very early stage of apoptosis, and preincubation of thymocytes with the inhibitors before dexamethasone or teniposide VM-26 were added irreversibly suppressed ladder formation. This implied that the cellular effector(s) of these compounds preexisted and were not resynthesized in response to the inducers of apoptosis. Serine protease inhibitors also suppressed apoptotic cell shrinkage and complete nuclear collapse, suggesting that these morphological changes were directly related to internucleosomal fragmentation of DNA. However, the serine protease inhibitors did not prevent high molecular weight DNA cleavage (> 50 kilobases) that preceded the ladder formation and thymocytes still died by apoptosis. This supported the view that internucleosomal DNA cleavage, considered to be the biochemical marker of apoptosis, might in fact be a late and dispensable step and that the newly described high molecular weight DNA cleavage might be a better indicator of apoptosis.Key words: serine protease, apoptosis, internucleosomal DNA fragmentation, high molecular weight DNA cleavage, protease inhibitors.

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