Multiple real-time fluorescent PCR detection kit for Y chromosome microdeletion

Abstract

The invention relates to the chromosome deletion detection field, and especially relates to a multiple real-time fluorescent PCR detection kit for Y chromosome microdeletion. The kit detects gene deletion at sY84 and sY86 loci of a Y chromosome AZFa subregion, sY127 and sY134 loci of a Y chromosome AZFb subregion, and sY254 and sY255 loci of a Y chromosome AZFc subregion by real-time fluorescent PCR. The kit realizes a stable and high-efficient multiple PCR reaction system by using homologous tailed primers and universal primers, adopts nucleic acid probes labeled by different fluorescent genes to indicate the presence of target fragments, and thus realizes the fastest simultaneous detection of sequences on 6 AZF deletion hotspots and 2 internally controlled genes by two tubes. The kit of the invention can be used for detection of male Y chromosome microdeletion genes, and has the advantages of high speed, accuracy, high throughput, low cost, no pollution, and the like.

Description

technical field [0001] The invention relates to the field of chromosome deletion detection, in particular to a multiple real-time fluorescent PCR detection kit and detection method for male Y chromosome microdeletion. Background technique [0002] Worldwide, about 15% of couples of childbearing age suffer from infertility, of which male factors account for about 50%. Studies have shown that more than 35% are due to genetic factors, mainly Klinefelter syndrome and Y chromosome microdeletion. It is currently known that there is azoospermia factor (AZF) on the Y chromosome, which is closely related to the production of sperm. There are three AZF subregions on the long arm of the Y chromosome—AZFa region, AZFb region, and AZFc region. Gene microdeletions in these regions can cause spermatogenesis disorders, oligospermia, weak sperm, or even azoospermia, resulting in male infertility. [0003] In recent years, assisted reproductive technology has developed rapidly, and intracy...

AI Technical Summary

Problems solved by technology

However, due to the non-sequence specificity of the dye, only singleplex PCR can be performed, which not only reduces the throughput, but also must rely on melting curve analysis to distinguish target fragments from primer-dimers

The current probe method is also single-plex PCR, which not only reduces the throughput, but also easily causes false positive results because there is no internal reference

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