| Literature DB >> 27595911 |
Jeremy D Henson1, Loretta M Lau2, Sylvia Koch3, Nancy Martin La Rotta3, Rebecca A Dagg4, Roger R Reddel2.
Abstract
The C-Circle Assay has satisfied the need for a rapid, robust and quantitative ALT assay that responds quickly to changes in ALT activity. The C-Circle Assay involves (i) extraction or simple preparation (Quick C-Circle Preparation) of the cell's DNA, which includes C-Circles (ii) amplification of the self-primed C-Circles with a rolling circle amplification reaction and (iii) sequence specific detection of the amplification products by native telomeric DNA dot blot or telomeric qPCR. Here we detail the protocols and considerations required to perform the C-Circle Assay and its controls, which include exonuclease removal of linear telomeric DNA, production of the synthetic C-Circle C96 and modulation of ALT activity by γ-irradiation.Keywords: ALT; C-Circle Assay; Exonuclease I; Native telomeric DNA dot blot; Quick C-Circle Preparation; Rolling circle amplification; Synthetic C-Circle C96; Telomere; Telomere maintenance mechanism; Telomeric qPCR; γ-Irradiation; λ exonuclease
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Year: 2016 PMID： 27595911 DOI： 10.1016/j.ymeth.2016.08.016
Source DB: PubMed Journal: Methods ISSN： 1046-2023 Impact factor: 3.608