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Application of reagent for detecting male infertility genes, kit for diagnosing male infertility and sequencing method

A male infertility, kit technology, applied in biochemical equipment and methods, microbial determination/examination, DNA/RNA fragments, etc., can solve problems such as inability to meet the needs of diagnosis, and achieve the effect of improving accuracy and sensitivity

Inactive Publication Date: 2020-04-07
GUANGDONG HYBRIBIO BIOTECH CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The diagnosis of typical clinical symptoms of male infertility (such as absence of vas deferens) can be detected by conventional methods, but for unexplained infertility (about 40% of infertile patients have abnormal spermatogenesis and no cause can be found), routine semen detection and Biochemical analysis can no longer meet the needs of its diagnosis

Method used

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  • Application of reagent for detecting male infertility genes, kit for diagnosing male infertility and sequencing method
  • Application of reagent for detecting male infertility genes, kit for diagnosing male infertility and sequencing method
  • Application of reagent for detecting male infertility genes, kit for diagnosing male infertility and sequencing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] The present embodiment provides a kit for auxiliary diagnosis of male infertility, which includes:

[0068] Library Concentration Detection Reagent: dsDNA HS reagent Kit; library preparation reagent: Ion AmpliSeq TM libraryKit; purification reagents: XP Reagent Kit; template-positive ISPs sequencing beads preparation reagent: Ion PGM TM Hi-Q TM 200Kit; water-in-oil PCR reagent: Ion PGM TM Hi-Q TM OT2 Kit; enrichment reagents: MyOne TM Streptavidin C1 Beads; Reagents for sequencing: Ion PGM TM Hi-QTM Sequencing Kit; and the first nucleic acid group for detecting pathogenic genes in male infertility patients and the second nucleic acid group for detecting microdeletions in the AZF region of Y chromosome in male infertility patients.

[0069] Wherein, the first nucleic acid group includes a primer pair A for detecting the pathogenic gene of the male infertility patient (as shown in Table 1), and the primer pair A includes primers whose sequences sequentially det...

Embodiment 2

[0072] Using the kit provided in Example 1, the samples to be tested were detected by high-throughput sequencing technology.

[0073] 1. Library construction.

[0074] The sample DNA to be tested is cut into fragments containing hundreds of bases, and then two different Ion Torrent adapters are connected to both ends of the DNA fragments to form a DNA sample library with two different adapters. After gap repair, length screening and other steps, a DNA fragment (library) of a certain length is obtained, the library is purified, and the library is amplified using primer pairs (Thermo Fisher Library Construction Kit Universal Library Construction Amplification System). Primers included primer pairs 1-398 provided in Example 1.

[0075] use 2.0 Fluorometer to analyze the quality of DNA in the solution, dilute the library to 100pM [275bp amplicon (blood sample) concentration is 22ng / mL].

[0076] 2. Template Preparation

[0077] The double-stranded DNA fragment is fixed on the...

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Abstract

The invention discloses application of a reagent for detecting male sterility genes, a kit for diagnosing male sterility and a sequencing method, and relates to the technical field of biology. Specifically, the reagent for detecting male sterility genes comprises STS sites of a plurality of AZF subregions and male sterility genes on autosomes and X chromosomes, can obtain reasons or related information of spermatogenic failure, and remarkably improves the accuracy and sensitivity of male sterility diagnosis.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of a reagent for detecting male infertility genes, a kit for diagnosing male infertility and a sequencing method. Background technique [0002] There is an azoosperm factor on the Y chromosome, which affects sperm production, and because the gene locus is too small to be judged by conventional methods, it is called Y chromosome microdeletion. [0003] The Y chromosome is the shortest apex centromere chromosome and is a male-specific chromosome. There are multiple genes involved in the spermatogenesis process on the Y chromosome. Studies have shown that there is a key fertility-related factor region on the Y chromosome, called Azoospermia factor (azoospermiafactor, AZF), the deletion of sites in the AZF region can cause patients to manifest as oligozoospermia or azoospermia, resulting in infertility. Moderate oligospermia and normal sperm count but abnormal morphology ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/6883C12Q2600/156
Inventor 刘翔郑焱李红霞陈微谢龙旭
Owner GUANGDONG HYBRIBIO BIOTECH CO LTD
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