DLO Hi-C (Digestion-Ligation-Only Hi-C) chromosome conformation capture method

Abstract

The invention discloses a DLO Hi-C (Digestion-Ligation-Only Hi-C) chromosome conformation capture method. The technology (the method) can overcome a series of shortcomings in a conventional chromosome conformation capture technology (Hi-C) which is loud in noise, high in cost, complex in experimental process, low in success rate, high in data analysis difficulty and the like, and the technology can conduct experiments just by simple ligation and digestion. The method provided by the invention has the following innovation points: 1) by conducting double cross-linking on target cells by virtue of EGS (ethylene glycol bis(succinimidyl succinate)) and a formaldehyde, the occurrence of decrosslinking in a later experimental process is prevented; 2) the application of biotin labeling is avoided in an experimental process, so that cost is reduced to a great extent; 3) a short time is consumed, and a library can be constructed within two and a half days just by simple ligation and digestion steps; 4) data noise is reduced, target gene interaction fragments are selectively recovered according to the sizes of the fragments by virtue of a page gel, and basically all data, obtained from sequencing is valid data; and 5) a library quality assessment standard is proposed for the first time, so that the quality of the library can be judged before conducting high throughput sequencing.

Description

technical field [0001] The invention relates to the field of chromosome conformation capture, in particular to a DLO Hi-C chromosome conformation capture method. Background technique [0002] From the start of genomics to today, the development of genomics has experienced two major waves. The first wave of development is represented by the Human Genome Project. From the early 1990s to 2003, the project sequenced the human genome, defined the main genes and their linear structures in the human genome, and greatly promoted The development of sequencing technology has opened up the era of genomics; the second wave is represented by the "Encyclopedia of Human Genome DNA Elements Project", which has systematically interpreted and annotated the human genome sequence since 2003. A large number of transcribed sequences and gene regulatory elements were discovered, and the expression and chromatin state of genes were defined. [0003] With the genome sequence, regulatory elements a...

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Problems solved by technology

[0003] With the genome sequence, regulatory elements and related annotations, researchers found that these discrete regulatory elements cannot effectively explain the regulatory mechanism and structure of many genes

Although the genomes of organisms store genetic information in their linear sequences, the correct expression and regulation of genes and the interaction between gene regulatory elements all need to be completed in the folding of chromosomes into complex three-dimensional structures, so scientists are faced with an urgent Problems to be solved: how chromosomes in the genome are folded, how linearly distant regulatory elements on the genome interact

[0008] 1) The cost is high, requiring biotin labeling and selective adsorption of streptavidin magnetic beads, which is very expensive and cannot be afforded by ordinary laboratories;

[0009] 2) The noise is high, and the streptavidin magnetic beads will adsorb a lot of impurity DNA fragments, resulting in too low useful data in the sequencing results;

[0010] 3) The test process is cumbersome, the cycle is long, and the success rate is low, and it often takes several tests to obtain usable test results;

[0011] 4) The quality of the library cannot be tested, and the quality of the library can only be analyzed after the sequencing results come out, and the risk of the experiment is very high;

[0012] 5) The analysis is difficult. It is necessary to remove the noise from the massive data, which causes great difficulty to the bioinformatics analysis.

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