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Enrichment of oligo(dG).oligo(dC)-containing fragments from human genomic DNA by Mg 2+-dependent triplex affinity capture.

Abstract：

:Oligo(dG).oligo(dC)- or short poly(dG).poly(dC)-containing fragments were enriched and cloned by means of Mg2+-dependent triplex affinity capture and subsequent cloning procedures. A library constructed after three cycles of enrichment showed that approximately 80% of the clones in the supercoiled form formed a complex with labeled oligonucleotide (dG)34. However, while the rest of the clones retained the ability to form a complex (type I clones), 90.9% failed to form a complex when they were linearized. This group of DNA was abundant in the genomic DNA, although it showed only approximately 3-fold enrichment by one cycle of affinity capture. This group was further classified into two species (types II and III) based on complex formation ability after phenol extraction. Type II clones retained the complex formation ability after treatment, while the human telomere [(TTAGGG)n] and telomere-like [(TGGAA)n] or [(TGGAG)n] sequences belonging to type III clones did not. Serial deletion experiments and the binding assays using oligonucleotides confirmed that the repetitive units containing T(G)nT ( n = 3-5) tracts or (G)n-motifs (n >/= 3) were the sites of complex formation for type II and III clones. On the other hand, type I clones contained poly(dG).poly(dC) tracts at least 10 nt long, and DNase I-footprinting analysis indicated that these tracts were the sites of complex formation.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Nishikawa N,Kanda N,Oishi M,Kiyama R

doi

10.1093/nar/25.9.1701

subject

Has Abstract

pub_date

1997-05-01 00:00:00

pages

1701-8

issue

9

eissn

0305-1048

issn

1362-4962

pii

gka306

journal_volume

25

pub_type

杂志文章

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