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Multi-omics sequencing and analyzing method based on Nanopore sequencing technology

An analysis method and sequencing technology, applied in the field of omics, can solve problems such as differences

Inactive Publication Date: 2020-07-28
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Application Information

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Problems solved by technology

The disadvantage of this calculation method is that it is susceptible to the influence of genes with extremely high expression and differential expression in different samples. The opening or closing of genes will affect the total number of molecules in the cell, which may lead to no significant difference after normalization of genes. difference, but there is a difference after normalization of genes that had no difference

Method used

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  • Multi-omics sequencing and analyzing method based on Nanopore sequencing technology
  • Multi-omics sequencing and analyzing method based on Nanopore sequencing technology
  • Multi-omics sequencing and analyzing method based on Nanopore sequencing technology

Examples

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Embodiment 1

[0097] Example 1 Multi-omics sequencing and analysis method based on Nanopore sequencing technology

[0098] E. coli strain K12 (WT wild strain) and EC135 strain (KO knockout strain, see ZL201210080124.6) with deletion of six endogenous restriction modification systems were used as experimental objects, and the growth curve was drawn by drawing the growth curve ( figure 1 ), select 4h during the exponential growth period and 13h during the plateau period to perform the following operations respectively.

[0099] 1. On the basis of maintaining the original ratio of nucleic acid in the sample, DNA and RNA were simultaneously extracted from samples of WT strain and KO strain in two periods (each with 3 biological replicates, a total of 12 samples). The kit used is Bacterial DNA / RNA / Protein kit (Qiagen, catalog number: 47054).

[0100] 2. Add Poly U tails to RNA samples

[0101] The 3' end of total RNA was labeled with E.coli PolyU polymerase (NEB technology, catalog number: ...

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Abstract

The invention provides a multi-omics sequencing and analyzing method based on a Nanopore sequencing technology. The method comprises the following steps: A, simultaneously performing DNA and RNA extraction on a collected sample; B, adding a Poly U tail to the 3' end of the extracted RNA; C, performing reverse transcription on the RNA with the Poly U tail to obtain cDNA, and adding an MID barcode sequence to the 5' end of the cDNA; D, mixing the cDNA sample with a DNA sample to build a library, and performing Nanopore sequencing; E, homogenizing the expression levels of all cDNA in sequencing data according to DNA; and F, reading and analyzing whole genome methylation signals in the sequencing data. According to the method, transcriptome and apparent methylation group detection can be realized on the basis of genome sequencing, and the purpose of multi-omics correlation analysis of a macro transcriptome, a macro genome and an apparent methylation group is achieved.

Description

technical field [0001] The present invention relates to the field of omics, in particular to multi-omics sequencing and analysis methods based on Nanopore sequencing technology. Background technique [0002] The host is actually a super ecosystem composed of the host itself and the symbiotic flora. A complex community of symbiotic flora lives on the host's skin surface, respiratory system, and reproductive system. The number and types of flora in the intestinal system Taking humans as an example, multiple studies have shown that there are as many as several hundred species of bacteria in the gut, and the total number of cells exceeds that of humans, and the genetic diversity contained in these different bacteria far exceeds that of humans. Humanity. A series of studies have shown that the flora plays an important role in the occurrence of health and disease. The occurrence of various chronic metabolic diseases, immune system diseases, liver diseases, central nervous system ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6869C12Q1/6806
CPCC12Q1/6806C12Q1/6869C12Q2563/143C12Q2563/149
Inventor 王军赵娜陈亮徐嘉悦于莹李亦嘉徐西占许亚昆
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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