Genome cytosine site epigenome typing method

A typing method and genomic technology, applied in the field of molecular biology, can solve problems such as epigenetic typing and typing, and achieve the effects of easy operation, popularization and application, mature technology, and low cost

Active Publication Date: 2017-06-13
BEIJING FORESTRY UNIVERSITY
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0004] At present, the analysis of methylation sequencing data involves the analysis of basic characteristics such as genomic methylation level, methylation distribution type and distribution tendency, but epigenetic typing is not yet possible

Method used

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  • Genome cytosine site epigenome typing method

Examples

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Effect test

Embodiment 1

[0039] Taking humans as the research object, the samples including parents and their offspring were subjected to bisulfate whole-genome methylation sequencing to obtain genome sequences;

[0040] 1) Genomic DNA extraction from the sample, using the CTAB method; 2) DNA sample detection, using agarose gel electrophoresis to analyze the degree of DNA degradation and whether there is RNA contamination, the detection reference figure 1 and figure 2 , figure 1 The medium sample is a qualified sample with the following electrophoresis bands: no degradation, no RNA pollution, figure 2Middle sample No. 4 was seriously degraded; No. 5 had severe RNA pollution. Then use Nanodrop to detect DNA purity (OD260 / 280), and finally use Qubit to accurately quantify the DNA concentration; 3) Library construction, when the sample is detected as A or B, it is considered qualified, and the addition ratio after passing is 1 / of the initial amount of library construction For the 1000 negative cont...

Embodiment 2

[0045] Taking Populus euphratica as the research object, the samples including the parents and their offspring were subjected to bisulfate whole-genome methylation sequencing to obtain the genome sequence;

[0046] 1) Genomic DNA extraction from the sample, using the CTAB method; 2) DNA sample detection, using agarose gel electrophoresis to analyze the degree of DNA degradation and whether there is RNA contamination, the detection reference figure 1 and figure 2 , figure 1 The medium sample is a qualified sample with the following electrophoresis bands: no degradation, no RNA pollution, figure 2 Middle sample No. 4 was seriously degraded; No. 5 had severe RNA pollution. Then use Nanodrop to detect DNA purity (OD260 / 280), and finally use Qubit to accurately quantify the DNA concentration; 3) Library construction, when the sample is detected as A or B, it is considered qualified, and the addition ratio after passing is 1 / of the initial amount of library construction For th...

Embodiment 3

[0051] Taking Populus microphylla as the research object, the samples including the parents and their offspring were subjected to bisulfate whole-genome methylation sequencing to obtain the genome sequence;

[0052] 1) Genomic DNA extraction from the sample, using the CTAB method; 2) DNA sample detection, using agarose gel electrophoresis to analyze the degree of DNA degradation and whether there is RNA contamination, the detection reference figure 1 and figure 2 , figure 1 The medium sample is a qualified sample with the following electrophoresis bands: no degradation, no RNA pollution, figure 2 Middle sample No. 4 was seriously degraded; No. 5 had severe RNA pollution. Then use Nanodrop to detect DNA purity (OD260 / 280), and finally use Qubit to accurately quantify the DNA concentration; 3) Library construction, when the sample is detected as A or B, it is considered qualified, and the addition ratio after passing is 1 / of the initial amount of library construction For t...

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Abstract

The invention provides a genome cytosine site epigenome typing method which includes the steps: 1) performing bisulfite whole genome methylation sequencing on male and female parents of samples to be tested and offspring samples to obtain male and female parent and offspring sample genome sequences; 2) comparing the male and female parent and offspring sample genome sequences with a reference genome to obtain comparison results to determine a cytosine site to be tested; 3) performing chromosome coordinate sequencing and reads replication removal on the comparison results, performing Call SNPs on 5-10bp upstream and downstream sequences of a known cytosine site through GATK2-V3.2 and accordingly distinguishing offspring allelic gene sequences; 4) comparing the obtained distinguished offspring allelic gene sequences with the male and female parent genome sequences, and finishing cytosine epigenome typing. The method finishes genome cytosine site epigenome typing for the first time and is mature in technology, low in cost and easy to operate, popularize and apply.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a genomic cytosine site epigenotype typing method. Background technique [0002] In higher eukaryotes, DNA methylation occurs only on the C on the G5′ side of the CpG dinucleotide. When the CpG-rich sequence (CpG island) is located in the gene promoter region, this modification has an important regulatory effect on the expression of the gene. In addition, it is also closely related to genome imprinting, gene inactivation of female X chromosome, cell proliferation, differentiation and development, tumor occurrence and development, and genetic instability. [0003] In recent years, methylation sequencing technology has gradually developed and improved, including bisulfate treatment of genomic methylation sequencing, using bisulfate treatment of genomic DNA to deaminate unmethylated cytosine into uracil, and methylation occurs. Kylated cytosine does not change. Methylat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G06F19/22C12Q1/68
CPCC12Q1/6858G16B30/00C12Q2525/191C12Q2523/125C12Q2531/113C12Q2539/113
Inventor 张德强
Owner BEIJING FORESTRY UNIVERSITY
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