Method for eliminating three-dimensional genomics technology noise by utilizing exonuclease combination

A genomics and exonuclease technology, applied in special data processing applications, instruments, electronic digital data processing, etc., can solve the problems of increasing technical efficiency and result specificity, so as to increase technical efficiency and result specificity, and improve production Out of proportion, the effect of simplification and elimination

Pending Publication Date: 2018-11-13
AGRI GENOMICS INST CHINESE ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, no report has paid attention to the high noise caused by this key connection step. Removing these linear DNA noises is an important focus for eliminating high noise, increasing technical efficiency and result specificity in 3D genomics technology (C-technology)

Method used

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  • Method for eliminating three-dimensional genomics technology noise by utilizing exonuclease combination
  • Method for eliminating three-dimensional genomics technology noise by utilizing exonuclease combination
  • Method for eliminating three-dimensional genomics technology noise by utilizing exonuclease combination

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1. Evaluation of the combined effect of exonucleases and their application in eliminating linear plasmid DNA.

[0029] 1. Selection of exonuclease and exonuclease combination

[0030] The following exonucleases were selected from NEB (New England Biolabs), as shown in Table 1.

[0031] Table 1 Exonucleases and their characteristics related to linear removal experiments

[0032]

[0033]A single exonuclease has a single function. Combining two exonucleases with complementary functions will have more comprehensive functions and better removal effects than a single exonuclease. According to functional complementarity and related literature reports, Lambda and RecJF exonuclease combination (see image 3 ): LRL combination, Lambda and RecJF exonuclease combined and made to react in Lambdabuffer; LRC combination, Lambda and RecJF exonuclease combined and made to react in CutSmart buffer.

[0034] 2. Exonuclease Combination Elimination of Linear Plasmid DNA Evalu...

Embodiment 2

[0041] Example 2. Application of exonuclease combination in noise elimination in key steps of C-technology.

[0042] C-techniques are used for three-dimensional genomics chromatin conformation grasping, which is a critical step derived from proximity junctions. How to increase the ratio of circularized junction molecules is very important for the subsequent detection of DNA ligation efficiency. C-technique prior to the adjacent connection step, the major steps are identical (see figure 2 ). In this example, the combination of exonucleases verified in the plasmid test in Example 1 was used to eliminate the linear "noise" DNA existing after the adjacent connection in the C-technology, and its efficient elimination effect was successfully verified. In the current C-technology, the starting amount of cells used is from 0.01 to 25 million. In the experiment, it was found that about 300 ng of DNA library can be obtained from 0.1 million starting cells, and about 30 ng of DNA libra...

Embodiment 3

[0049] Example 3. Application of Exonuclease Combination in Hi-C Technology for Noise Elimination.

[0050] Hi-C is a high-throughput chromatin conformation capture technology based on 3C technology, which can capture the entire genome interaction mediated by proteosomes. Proximity connectivity is a key step in Hi-C technology. Proximity connections are currently noisy, and this step is one of the main reasons for the low effective interaction rate of standard Hi-C (see figure 1 and figure 2 ). Example 1 The exonuclease combination elimination method has been verified in the plasmid test. Example 2 The combined elimination method of exonucleases has been validated for noise reduction after the common core step of C-technique—“adjacent joining”. This example intends to demonstrate the elimination effect of the exonuclease combination noise elimination method applied in one of the specific C-technical members—Hi-C technology: increase the effective interaction ratio.

[00...

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Abstract

The invention discloses a method for eliminating three-dimensional genomics technology noise by utilizing exonuclease combination. The method comprises the step of adding exonuclease combination to eliminate adjacently connected linear DNA when performing the adjacent connection at the key step of the three-dimensional genomics technology, thereby improving the cyclic effective interaction proportion. The excellent performance of the exonuclease combination for eliminating the linear DNA provides an effective measure for eliminating the noise through the enzyme digestion of the exonuclease combination after the C-technology connection; the method for eliminating the three-dimensional genomics technology noise by utilizing exonuclease combination has the advantages of being convenient, fastand efficient, the possibility of producing the false-positive product or signal can be reduced, the effective interaction cyclic DNA proportion is enhanced, the terminal effective interaction data output ratio is greatly improved, the sequencing space is saved, the sequencing cost is reduced, and the operation for eliminating the resource consumed by ineffective information in the big data by using the complex biological information at later is simplified.

Description

technical field [0001] The invention relates to the technical field of genome sequencing, in particular to three-dimensional genomics, and relates to a method and application for eliminating noise of three-dimensional genomics technology by using exonuclease combination. Background technique [0002] Chromatin conformation capture technology (C-technology) is an important technology in 3D genomics technology. C-technology is widely used to elucidate the genome interaction folding mode, reconstruct the three-dimensional topology of the genome, and study important biological issues such as transcriptional regulation and cell fate determination of organisms. C-technology, mainly developed from 3C technology (chromatin conformation capture technology), including 3C, 4C (circularized chromatin conformation capture technology), 5C (chromatin conformation capture carbon copy technology), Hi-C (Qualcomm Quantitative chromatin conformation capture sequencing technology), ChIA-PET (c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G06F19/18G06F19/20
Inventor 张玉波孔思远张高林黄其通范磊李清黄雷彭艳玲
Owner AGRI GENOMICS INST CHINESE ACADEMY OF AGRI SCI
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