Primer composition, kit and method for detecting the expression level of insulin-like peptide ilp series genes in Drosophila melanogaster

Abstract

The invention discloses a primer composition, a kit and a use method for detecting the expression level of insulin-like peptide ILP series genes in Drosophila melanogaster. The primer composition comprises RT amplification primers and PCR amplification primers of insulin-like peptide and RNA internal reference. The kit includes DEPC water, 5×RT buffer, reverse transcription primer, reverse transcriptase, Z solution, 10×PCR buffer, PCR primer, 25mM magnesium chloride solution, DNA polymerase and positive control. The present invention can detect 7 insulin-like peptides of Drosophila melanogaster at the same time, and can complete the detection of 192 samples in one day, which not only saves production and detection costs, but also improves detection efficiency; the RNA internal reference provides a control internal reference for the integrity of the sample RNA, Ensure the judgment of sample quality during the inspection process, avoid false negatives, and make the detection have better sensitivity and specificity, thereby avoiding the problem of low specificity of other detection methods.

Description

technical field [0001] The invention relates to a multiple gene detection kit and its application, in particular to a primer composition, a kit and a use method for detecting the expression level of insulin-like peptide ILP series genes in Drosophila melanogaster. Background technique [0002] In humans and mammals, insulin is an important hormone secreted by the beta cells of the pancreas. Insulin is the only hormone that lowers blood sugar in the body, and it is also the only hormone that simultaneously promotes the synthesis of glycogen, fat, and protein. Insulin can also promote the entry of potassium ions and magnesium ions through the cell membrane into the cell; and promote the synthesis of deoxyribonucleic acid (DNA), ribonucleic acid (RNA) and adenosine triphosphate (ATP). Due to the important role of insulin in metabolic regulation, the role of insulin signaling pathway in various physiological and pathological processes (eg, cancer, aging) has received increasing...

AI Technical Summary

Problems solved by technology

The advantages of fluorescent quantitative PCR technology: high sensitivity and accurate quantification, but there are also the following disadvantages: 1) low throughput: if only one fluorescent marker is used, only one gene can be detected at a time

When a sample needs to detect multiple genes at the same time, it must be tested one by one, the cost is relatively high, the efficiency is low, the cycle is long, and it is impossible to start with a large number of samples

2) The cost is relatively high: if multiple genes need to be detected at the same time, multiple fluorescent markers are required; at present, a maximum of 4 to 5 fluorescent markers are used in common to mark each gene segment to be detected, so multiple fluorescent markers are used relatively high cost

[0006] (2) The fluorescent in situ hybridization method is to hybridize fluorescently labeled gene probes to intracellular nucleic acids in situ, and detect the brightness and quantitative gene expression levels through a fluorescent microscope, which has the disadvantages of low throughput, low sensitivity, and high cost

It has the advantages of high throughput, but the technology is expensive and complicated, and the synthesis and fixation of probes are complicated, and it cannot be accurately quantified, with poor repeatability and low sensitivity.

[0008] (4) As a relatively new technology, RNA sequencing has the advantages of high sensitivity and high throughput, but it is expensive

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