Serum miRNA combination as molecular marker for assessing non-obstructive azoospermia

A molecular marker, non-blocking technology, applied in the direction of DNA/RNA fragment, recombinant DNA technology, microbial determination/inspection, etc., can solve the problems of weak correlation of fertilization ability, difficult standardization of semen, etc., to achieve high accuracy, Simple to use effects

Inactive Publication Date: 2019-12-20
广东省计划生育科学技术研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although great achievements have been made in the assessment of semen quality, the detection of semen is still difficult to standardize
Some...

Method used

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  • Serum miRNA combination as molecular marker for assessing non-obstructive azoospermia
  • Serum miRNA combination as molecular marker for assessing non-obstructive azoospermia
  • Serum miRNA combination as molecular marker for assessing non-obstructive azoospermia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Microarray detection of differential expression of miRNA in the serum of patients with non-obstructive azoospermia and healthy sperm donors

[0029] 1. Experimental group

[0030] Sample group: Serum samples from patients with non-obstructive azoospermia were obtained from Guangdong Provincial Family Planning Specialized Hospital, and were diagnosed as patients with non-obstructive azoospermia by the Andrology Department of our hospital;

[0031] Control group: Serum samples from healthy males who donated sperm to Guangdong Human Sperm Bank and met the conditions for donating sperm. The diagnostic criteria of non-obstructive azoospermia are azoospermia with no sperm found in two routine semen analyzes except for vas deferens obstruction factors.

[0032] 2. Experimental operation

[0033] The samples were collected with the consent of the Ethics Committee of the Guangdong Provincial Family Planning Specialty and after the patients’ informed consent.

[0034...

Embodiment 2

[0044] Example 2 Real-time quantitative PCR to verify the differential expression of miRNA in the serum of patients with non-obstructive azoospermia and healthy sperm donors

[0045]According to the results of the miRNA microarray chip, five miRNAs with higher Fold change (hsa-miR-1263, hsa-miR-221-5p, hsa-miR-483-3p, hsa-miR-4275 and hsa-miR-194 -3p), using Mir-X TM miRNAqRT-PCR Kit for real-time quantitative PCR validation.

[0046] 1. Experimental operation

[0047] RNA was extracted and reverse-transcribed using Mir-X miRNA First-Strand Synthesis Kit. The primer for reverse transcription was: 5-GCTGTCAACGATACGCTACGTAACGGCATGACAGTG-3, and 5SrRNA was used as an internal reference gene. Others were carried out according to the instructions of the kit. The upstream primers used are listed in Table 2. The downstream primer used the 3' primer in the kit: 5-GCTG TCAACGATA CGCTACG TAA-3'.

[0048] The reaction was amplified on a 96-well STEPONEPLUS (ABI, USA) fluorescent P...

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Abstract

The invention discloses an application of a serum miRNA combination as molecular marker for assessing non-obstructive azoospermia. The serum miRNA combination includes hsa-miR-1263, hsa-miR-221-5p, hsa-miR-483-3p, hsa-miR-4275 and hsa-miR-194-3p. Through the miRNA combination, a reagent kit for assessing the non-obstructive azoospermia is prepared. The serum miRNA for assessing the non-obstructiveazoospermia is obtained, further a detection reagent kit containing primers used for quantitative RQ-PCR detection of the serum miRNA combination is obtained, detection is performed through the detection primers and the detection reagent kit, the operation is simple, the accuracy is high, and the serum miRNA combination is worth of extensive popularization.

Description

technical field [0001] The invention relates to the technical field of medical detection, and more specifically relates to a serum miRNA combination used as a molecular marker to evaluate non-obstructive azoospermia. Background technique [0002] At present, about 10%-15% of couples of childbearing age in the world are infertile, and 50% of them are related to male factors. Non-obstructive azoospermia (NOA) is the most common type of male infertility, with an incidence of about 1% in adult males. In addition to infection, endocrine, immunity, heredity, congenital malformations and varicocele and other etiologies, 50% of male infertility has no clear etiology and is diagnosed as idiopathic infertility. It is generally believed that idiopathic infertility is associated with unknown genetic and epigenetic factors. The etiology of male infertility is complex, but abnormal semen is one of the most common laboratory indicators. Semen quality is directly related to the fertiliza...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/6883C12Q2600/158C12Q2600/178
Inventor 赵文忠傅文婷吴瑞珊钟安周雨
Owner 广东省计划生育科学技术研究所
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