Noninvasive antenatal haplotype construction method based on long fragment DNA capturing and three-generation sequencing

Abstract

A noninvasive antenatal haplotype construction method based on long fragment DNA capturing and three-generation sequencing includes the steps of creating a second-generation library of genomes DNA inperipheral blood of pregnant women and / or husbands of the pregnant women; capturing target genes and flank regions; creating a third-generation computer sequencing library, and performing third-generation sequencing to acquire the sequencing read length; extending from mutational sites of target genes on the sequencing read length to two ends so as to look up heterozygous SNP sites; when differentsequencing read length overlapping regions contain one or more same SNP sites, continuing to differentiate the haplotype towards two ends successfully until a section of region is not covered by thesequencing read length or the SNP sites detected by the sequencing read length are homozygous. The noninvasive antenatal haplotype construction method has the advantages that haplotype construction ofparent individuals is realized, and the defect that current common noninvasive antenatal detection based on parent-child family haplotype analysis is not applicable to family detection without acquisition of propositus samples is overcome.

Description

technical field [0001] The invention relates to the technical field of sequencing, in particular to a non-invasive prenatal haplotype construction method based on long-fragment DNA capture and three-generation sequencing. Background technique [0002] According to the World Health Organization's 2015 birth defect report, there are about 3 genes related to birth defects for every 100 newborns in the world. Every year, 3.2 million newborns with birth defects are born, and 270,000 newborns die of birth defects. Studies have shown that the vast majority of birth defects are related to genetic factors, and single-gene defects are one of the important factors. At present, there are no radical measures for most single-gene defects, and only life-long replacement treatments are available. Most of them are permanently disabled or mentally handicapped, which cannot be cured, thus causing heavy economic and psychological burdens to society and families. Prenatal testing for high-risk ...

AI Technical Summary

Problems solved by technology

STR linkage analysis has the problem of fewer STR linkage marker sites. In specific cases, there may be no available STR sites, which requires a lot of pre-experimentation, and most of the STRs are far away from the defect site, so the possibility of misdiagnosis caused by recombination cannot be ruled out.

Based on haplotype analysis, the method of genome capture sequencing or SNP typing of the parental proband's family is often used to first obtain the haplotype associated with the defect site. The multiplex PCR operation is complicated, and the cost of family capture sequencing is high, and it is difficult to popularize. It is necessary to obtain the samples of the parent-child family at the same time, but in practical applications, it is usually encountered that the samples of the offspring of the couple to be tested are not available. For example, Lu Sijia disclosed "a method for SNP-haplotype analysis using multiplex PCR technology" (Publication number: CN105385755A)

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