Mir-19a detection kit based on allglo probe fluorescence quantitative PCR and detection method thereof

Abstract

miR-19a detection kit and detection method based on AllGlo probe fluorescent quantitative PCR. The detection kit is equipped with a box body, a partition, an exogenous reference bottle, a neck ring reverse transcription reagent bottle, and a real-time fluorescence quantitative PCR reagent bottle; the partition is set in the box body, and the exogenous reference bottle, neck ring reverse transcription reagent bottle , The real-time fluorescent quantitative PCR reagent bottle is inserted on the partition, the exogenous reference bottle is equipped with an exogenous reference, the neck ring reverse transcription reagent bottle is equipped with a neck ring reverse transcription reagent, and the real-time fluorescent quantitative PCR reagent bottle is equipped with a real-time fluorescent quantitative PCR reagents. Detection method: extract miRNA in the sample; use the stem-loop reverse transcription reagent provided by the detection kit to reverse miRNA into cDNA; use the real-time fluorescent quantitative PCR reagent provided by the detection kit to perform real-time fluorescent quantitative PCR amplification of cDNA; comprehensive analysis instrument Given the various data, set the threshold and baseline, and analyze the results.

Description

technical field [0001] The invention belongs to the fields of laboratory medicine, clinical medicine, biotechnology and molecular biology, and in particular relates to a miR-19a detection kit based on AllGlo probe fluorescence quantitative PCR and a detection method thereof. Background technique [0002] MicroRNA (miRNA) is a kind of non-coding RNA molecule of about 22 nucleotides widely present in eukaryotic cells, which participates in many physiological and pathological processes in organisms, by regulating gene expression, in cell proliferation, apoptosis , growth and development, cell differentiation, metabolism and other processes play an important role. The specific performance is that miRNA forms the initial primary transcript pri-miRNA to pre-miRNA in the nucleus, and then transports out of the nucleus, forms mature miRNA by shearing, and forms RISC (RNA-induced silencing complex) with Ago protein, etc. Partially inhibit or degrade the target mRNA sequence and part...

AI Technical Summary

Problems solved by technology

Among them, northern blot technology is one of the early means of RNA detection, but its specificity and sensitivity are low. If the sample RNA content is low or there is degradation, it may not be detected; in situ hybridization technology is used to detect the distribution of miRNA at the tissue and cell level , while performing semi-quantitative detection of miRNA, its disadvantage is that it cannot accurately detect low-expression miRNA; microarray chip technology is a high-throughput detection technology that can simultaneously detect a large number of miRNAs in one or more samples , but there are problems such as time-consuming and labor-intensive chip fabrication, high labeling costs, and low detection sensitivity and specificity.

The reverse transcription primer based on the stem-loop structure can specifically recognize and amplify the mature miRNA sequence, but the precursor of the miRNA has not been amplified. The disadvantage of the Taqman-MGB probe for detecting miRNA is that the synthesis is expensive, which is not conducive to popularization.

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