PCR detection method for human hsa-miR-124a-3p and application of PCR detection method to DS (Down's syndrome) pregnancy rick prediction

Abstract

The invention discloses a PCR detection method for human hsa-miR-124a-3p. The PCR detection method comprises the following steps: 1) extracting total RNA; 2) performing RT-qPCR detection: an RT primer is 5'-GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACGGCATTC-3', an RTreaction procedure refers to 16DEG C for 30min, 42DEG C for 30min and 85DEG C for 5min; qPCR procedure refers to 50DEG Cfor 2min, 95DEG C for 10min, 95DEG Cfor 15s, 60DEG C for 60s, and 35cycles; 3)calculating concentration of to-be-detected hsa-miR-124a-3p. The invention further discloses an application of the PCR detection method to DS (Down's syndrome) pregnancy rick prediction.10<-5>-10<-3> pmol / mu L is determined as a DS high risk interval value of the detection method.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to a human hsa-miR-124a-3p PCR detection method and its application in DS pregnancy risk prediction. Background technique [0002] Down's syndrome (Down's syndrome, DS) is the most common genetic disease in humans that causes mental retardation and developmental disorders in newborns, also known as trisomy 21 (trisomy 21, T21) or congenital stupidity Its incidence rate in newborn fetuses is about 1 / 700, while the incidence rate of DS in my country is about (1.53-2.40) / 10,000. Most fetuses with DS stop developing or cause miscarriage in utero. Therefore, the actual incidence of DS should be much higher than 1 / 700. And with the increase of women's pregnancy age, especially the change of my country's current population and childbearing policy, the increase of older or elderly pregnant women and the further promotion and application of assisted reproductive technol...

AI Technical Summary

Problems solved by technology

Down's screening is highly safe, but its specificity is poor. Although the prevalence of DS is as high as 84% ​​when the fetal nuchal translucency (NT) > 2.5 mm, it still cannot meet the requirements for diagnosis. Down's screening High-risk pregnant women still need follow-up examinations to confirm the diagnosis

Non-invasive DNA testing is a prenatal detection method for DS that has emerged in recent years, also known as non-invasive prenatal DNA testing or non-invasive fetal chromosome aneuploidy testing. And its sequence analysis is performed to obtain the genetic information of the fetus, so as to diagnose DS, Patau Syndrome (PS) and Edwards Syndrome (Edwards Syndrome, ES), its safety and sensitivity are high, but the test The cost is high, the technical operation is complicated, and the inspection cycle is long. At present, the competent health department implements qualification certification management for it, and it is mostly used in third-party medical testing institutions, and it is not easy to promote it in clinical practice, especially in grassroots medical institutions.

Amniocentesis, chorionic villus puncture, and fetal umbilical cord blood examination all use invasive procedures to obtain fetal exfoliated cells and conduct chromosome karyotype analysis on them to achieve the purpose of diagnosis. Patients have high clinical experience requirements, and there is a 1% chance of miscarriage

At present, although karyotype analysis is the gold standard for prenatal diagnosis of DS, its detection technology is complicated, the conditions are high and the detection cycle is long, so it is limited to high-risk pregnant women for Down's screening

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