Tagged Chromosomal Insertion Site System: A Method to Study Lamina-Associated Chromatin.

Abstract:

:The three-dimensional (3D) organization of the genome is important for chromatin regulation. This organization is nonrandom and appears to be tightly correlated with or regulated by chromatin state and scaffolding proteins. To understand how specific DNA and chromatin elements contribute to the functional organization of the genome, we developed a new tool-the tagged chromosomal insertion site (TCIS) system-to identify and study minimal DNA sequences that drive nuclear compartmentalization and applied this system to specifically study the role of cis elements in targeting DNA to the nuclear lamina. The TCIS system allows Cre-recombinase-mediated site-directed integration of any DNA fragment into a locus tagged with lacO arrays, thus enabling both functional molecular studies and positional analysis of the altered locus. This system can be used to study the minimal DNA sequences that target the nuclear periphery (or other nuclear compartments), allowing researchers to understand how genome-wide results obtained, for example, by DNA adenine methyltransferase identification, chromosome conformation capture (HiC), or related methods, connect to the actual organization of DNA and chromosomes at the single-cell level. Finally, TCIS allows one to test roles for specific proteins in chromatin reorganization and to determine how changes in nuclear environment affect chromatin state and gene regulation at a single locus.

journal_name

Methods Enzymol

journal_title

Methods in enzymology

authors

Harr JC,Reddy KL

doi

10.1016/bs.mie.2015.09.028

subject

Has Abstract

pub_date

2016-01-01 00:00:00

pages

433-53

eissn

0076-6879

issn

1557-7988

pii

S0076-6879(15)00546-7

journal_volume

569

pub_type

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