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Amplification composition for Y chromosome microdeletion detection and kit thereof

A Y chromosome and microdeletion technology, applied in the field of amplification compositions and kits for Y chromosome microdeletion detection, can solve problems such as increasing the difficulty of primer design, and achieve the effects of intuitive and accurate result interpretation, short detection time, and simple operation.

Active Publication Date: 2019-07-02
BEIJING HARMONY HEALTH MEDICAL DIAGNOSTICS CO LTD
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AI Technical Summary

Problems solved by technology

[0004] Multiplex PCR combined with agarose gel electrophoresis is currently the most widely used method for Y chromosome microdeletions. After years of clinical practice, this method has been proved to be accurate, sensitive, easy to operate, and low in cost. However, the biggest difficulty and challenge of this method lies in the multiple Design of PCR primers: (1) Accurately locate the position of the STS and design primers to avoid STS loss; (2) Design the primers scientifically and reasonably to avoid mutual interference; (3) The size difference between the amplified fragments must reach more than 50bp, agar Glycogel electrophoresis can be used to better distinguish and interpret the results
These limitations greatly increase the difficulty of primer design

Method used

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  • Amplification composition for Y chromosome microdeletion detection and kit thereof
  • Amplification composition for Y chromosome microdeletion detection and kit thereof
  • Amplification composition for Y chromosome microdeletion detection and kit thereof

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Embodiment Construction

[0063] A kit for detecting a Y chromosome microdeletion, which is detected by PCR amplification-agarose gel electrophoresis, comprising the following steps:

[0064] (1) Design of primer combination

[0065] Select 6 STS sites (AZFa: sY84, sY86; AZFb: sY127, sY134; AZFc: sY254, sY255) and 2 quality control sites (ZFX / ZFY, SRY) according to the recommendations of EAA / EMQN guidelines, and search through NCBI According to the DNA sequence of each site, design and synthesize the best amplification primers to ensure that the size difference between the amplified fragments is more than 77bp, and will not interact to produce non-specific products or dimers. Then, it was diluted to the required concentration of 10 μM as required, and the specific primer sequences are shown in Table 1, as shown in SEQ ID NO:1-16 in the sequence listing:

[0066] Table 1 Primer Sequence

[0067]

[0068] (2) DNA extraction

[0069] Genomic DNA was extracted from the collected fresh peripheral bloo...

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Abstract

The invention discloses an amplification composition for Y chromosome microdeletion detection and a kit thereof. By using primers for amplifying 6 STS sites (AZFa: sY84, sY86; AZFb: sY127, sY134; AZFc: sY254, sY255) and 2 control sites (ZFX / ZFY, SRY), and the six STS sites closely related to the Y chromosome microdeletion are detected by a multiplex PCR combined agarose gel electrophoresis technology to determine the Y chromosome microdeletion. With the amplification composition of the present invention, six STS sites can be simultaneously detected by only two tube amplification reactions.

Description

technical field [0001] The invention relates to the technical field of chromosome deletion detection, in particular to an amplification composition and a kit for detection of Y chromosome microdeletion. Background technique [0002] On the long arm of the Y chromosome, there is an area of ​​azoosperm factor (AZF) that affects spermatogenesis, which can be further divided into three areas: AZFa, AZFb and AZFc. Microdeletions in any one region can cause spermatogenesis disorders in men, leading to oligospermia or azoospermia. In male infertility patients caused by spermatogenesis disorders, the incidence of Y chromosome microdeletion is second only to Klinefelter's syndrome (Klinefelter's syndrome), which is the second genetic factor. Y chromosome microdeletion has become a routine examination item for male infertility patients. [0003] In the three regions of AZF, each region has a clear sequence tag site (STS), and the location of the deletion break has been identified. ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12Q1/6858
CPCC12Q1/6883C12Q1/6858C12Q2600/156C12Q2600/16C12Q2531/113C12Q2537/143C12Q2565/125
Inventor 智慧芳佟洪梅赵方圆倪君君
Owner BEIJING HARMONY HEALTH MEDICAL DIAGNOSTICS CO LTD
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