Purification of apoptotic DNA

LOCKSHIN, RICHARD A YPRLBIO at sjumusic.stjohns.edu
Thu Jun 9 14:58:49 EST 1994


In article <sabol.1121395919A at newssrv.dcrt.nih.gov> sabol at codon.nih.gov (Steven L. Sabol) writes:
>In apoptosis, genomic DNA is usually partially degraded, with a hallmark
>"nucleosomal ladder" of 190-200 bp and multiples thereof appearing, although
>in many systems only a minority of the genomic DNA is degraded this far,
>much of it remaining in 50+-kilobase fragments.
>
>To analyze apoptotic DNA, it would seem to be desirable to isolate genomic
>DNA of all sizes rather than just small ladder DNA.  Standard DNA isolation
>procedures accomplish this, but there are often problems in redissolving
>ethanol-precipitated high-molecular weight DNA without extensive shearing.
>Several companies sell genomic DNA purification kits involving ion-exchange
>columns and protocols that do not include ethanol precipitation.  Has anyone
>had experience with these kits in purifying apoptotic DNA?  If so, how does
>column-kit-purified DNA compare with DNA purified by non-column methods? How
>do the recoveries of ladder-size DNA vs. 50+-kilobase DNA compare in these
>column-based kits?
>
>Steven L. Sabol
>Lab. of Biochemical Genetics
>NHLBI, NIH
>Bethesda, MD
>sabol at codon.nih.gov
>.
I agree with you that some people "purify" DNA fragments to the point
that there is virtually nothing left other than artifactual fragmenta-
tion.  Check our article (Zakeri, Lockshin, et al, FASEB J spring of
1993) in which we compared purified DNA and total DNA by end-labeling.
Let me know what else you come up with.
Richard Lockshin, Dept. Biol. Sci. St. John's University Jamaica
NY 11439 718:  990-1854, FAX 718:  380-8543 yprlbio at sjumusic.stjohns.edu






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