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Quantitative test method of exopalaemon carinicaudamitochondrial genome copy number

A technology of mitochondrial genome and quantitative detection method, which is applied in the field of molecular biology to achieve high specificity, simple operation and rapid detection effect

Inactive Publication Date: 2017-03-15
HUAIHAI INST OF TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the detection method of mtDNA copy number in aquatic organisms has only been reported in a few fish, and there is no relevant research on marine crustaceans

Method used

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  • Quantitative test method of exopalaemon carinicaudamitochondrial genome copy number
  • Quantitative test method of exopalaemon carinicaudamitochondrial genome copy number

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Embodiment Construction

[0018] The present invention can be realized through following several operation steps:

[0019] 1. According to the nuclear genome (nDNA) single-copy gene (select two here: adenylate transferase gene (ANT) and trypsin gene (trypsin)) and mitochondrial genome (mtDNA) single-copy gene (select four : ATP6, ND1, COII and D-loop) sequences, design corresponding PCR amplification primers for each gene fragment, the fragment size is between 90-130bp. Through conventional PCR reaction, verify the amplification effect of the designed primers, and sequence the DNA fragment amplified by the primers to determine whether it is the target amplified region of the corresponding gene. According to the clarity of the amplified band and the specificity of the amplification, and In combination with the quantitative stability characteristics of these single-copy genes in the follow-up fluorescent real-time quantitative PCR reaction, the ATP6 gene in the adenylyltransferase gene (ANT) on the nucle...

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Abstract

The invention discloses a quantitative test method of exopalaemon carinicauda mitochondrial genome copy number. The method comprises the steps of selecting the adenylyl transferase gene (ANT) and the ATP6 respectively as the single copy genes of the nuclear genome and the mitochondrial genome of white tailed shrimp, designing the primer and the fluorescent probe suitable for the real time quantitative PCR reaction of the ANT and ATP6 genes, preparing real time quantitative PCR reaction standard of the ANT and ATP6 genes, through drawing the standard curve (as shown in the attachment), and using the standard curve as the standard, and quantifying and calculating the copy number of the mitochondrial genome in the to-be-tested sample cells. The method can correctly and quantitatively test the mitochondrial genome copy number of the exopalaemon carinicauda tissue sample cells, and has the advantages of being high in specificity, and simple in operation. Merely the fluorescence quantitative PCR instrument is enough to complete the quantification of the exopalaemon carinicauda mitochondrial genome copy number.

Description

[0001] Technical field: the invention belongs to the technical field of molecular biology, and relates to a quantitative detection method for the mitochondrial genome copy number of the white shrimp. [0002] Background technique: [0003] Existing studies have shown that mitochondria are very active organelles in biological life activities. During the process of growth and development, not only their shape will change to a certain extent, but also their number will change significantly. Essentially, at different stages of growth and development, organisms must face the process of cell proliferation and apoptosis, and this periodic activity of cells also regulates the activity of mitochondria in cells. The current research shows that when the cells of the organism are in different conditions, the copy number of the mitochondrial genome (mtDNA) will also change, and this change is very different. The mtDNA copy number in a cell can vary from several to thousands. This mainly de...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/686C12Q2561/113C12Q2563/107C12Q2545/114C12Q2531/113C12Q2545/113
Inventor 高焕张培赖晓芳李志辉薛蓓赵莲阎斌伦
Owner HUAIHAI INST OF TECH
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