Chromosome conformation analysis

摘要

Disclosed herein are compositions, methods and kits for analyzing three-dimensional chromatin and/or chromosome conformation. Method are also disclosed for using the methods disclosed herein for diagnosing diseases such as cancer.

主权项

1. A method of determining the three-dimensional arrangement of the chromatin within a cell, comprising:
a) isolating cells from a biological sample; b) incubating the cells with a cross-linking agent, thereby cross-linking proteins with DNA and forming a cross-linked product; c) lysing the cells; d) digesting the DNA with a restriction endonuclease; e) incubating the digested DNA with a phosphatase; f) ligating a first looped half-adaptor to the digested DNA thereby forming a first ligation product and a second looped half-adaptor to the digested DNA thereby forming a second ligation product, wherein the first and second looped half-adaptors are different from each other and each looped half-adaptor comprises a stem-loop structure complementary to the stem-loop structure of the other half-adaptor, wherein the 5′ end of the stem of each looped half-adaptor comprises a 5′-overhang that terminates in a 5′-phosphate group and is ligated to the digested DNA; g) heating the first and second ligation products thereby allowing the half-adaptors to anneal to each other; h) treating the product of step g) with a kinase and a ligase to ligate the looped half-adaptors to the digested DNA thereby forming a third ligation product; and i) analyzing the third ligation product.