Real-time quantification PCR chip used for detecting gene expression of mouse cholesterol metabolism

A real-time quantitative and gene expression technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve problems such as lack of availability, no targeted research on genes, and limited accuracy of read numbers

Inactive Publication Date: 2014-05-28
DONGHUA UNIV
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Problems solved by technology

However, these two methods have the following disadvantages: there is no accurate quantitative information in the expression profile chip, and the results need a large number of experimental verifications; the expression profile chip is a broad-spectrum screening method, and it does not study genes in a targeted manner. Unrelated genes will generate a lot of useless or wrong information and interfere with the analysis of results; RNA-Seq can detect all RNA in a sample, and a large part of RNA in cells comes from ribosomes and mitochondria, limiting the number of reads for other RNAs and the accuracy of these RNA expression levels
At the same time, these two methods have high research costs and can only be completed in professional experimental centers
Not eligible to be completed in a general laboratory

Method used

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  • Real-time quantification PCR chip used for detecting gene expression of mouse cholesterol metabolism
  • Real-time quantification PCR chip used for detecting gene expression of mouse cholesterol metabolism
  • Real-time quantification PCR chip used for detecting gene expression of mouse cholesterol metabolism

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Embodiment 1

[0063] In this study, by designing a stable and reliable qPCR array detection scheme, mouse hepatocellular carcinoma cell Hepa1-6 was used as the research object, and qPCR array was used to detect the mRNA expression of cholesterol metabolism genes in Hepa1-6 induced by high glucose, aiming to explore the mRNA level. Effects of high glucose on genes involved in cholesterol synthesis.

[0064] 1 Materials and reagents

[0065] Mouse hepatoma cells Hepa1-6 were purchased from the Cell Resource Center, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. Low DMEM medium (Hycolon, USA), fetal bovine serum (Hycolon, USA), D-glucose (Sigma, USA), Trizol (Invitrogen, USA), DNase1, reverse transcription reagent (Fermentas, USA), SYBR real-time PCR Premixture kit (Bioteke), PCR primers were designed and synthesized by Shanghai Sangon. Cell incubator (Thermo, USA), real-time fluorescent quantitative PCR instrument (ABI7500, USA).

[0066] 2 Cell culture, experime...

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Abstract

The invention relates to a real-time quantification PCR chip used for detecting gene expression of mouse cholesterol metabolism. The real-time quantification PCR chip uses 88 genes closely related to cholesterol metabolism as detection sites and selects 4 house-keeping genes. The real-time quantification PCR chip provided by the invention can carry out target analysis on gene related to mouse cholesterol metabolism, needs a short experimental period, has simple data processing and low cost, does not need special experiment organization detection, needs only a quantification PCR instrument and has the advantages of high amplification efficiency, reliable quantification results, good repeatability and high specificity.

Description

technical field [0001] The invention belongs to the field of detecting the expression of cholesterol genes, in particular to a real-time quantitative PCR chip (qPCR array) for detecting the expression of cholesterol metabolism genes in mice. Background technique [0002] Currently, techniques for studying differences in gene expression mainly include expression profiling microarrays, RNA sequencing, and qPCR arrays. The expression profile chip uses cDNA or oligonucleotide fragments as probes, which are solidified on the chip; the mRNA of the test sample (treatment group) and the control sample are labeled with two different fluorescent molecules, and then hybridized with the chip at the same time , to detect changes in gene expression levels by analyzing the ratio of the fluorescence intensities of two samples hybridized with the probe. RNA sequencing, also known as transcriptome sequencing. Transcriptome refers to the collection of all transcribed mRNA products of a speci...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6851C12Q2531/113C12Q2561/113C12Q2545/113
Inventor 朱长保肖君华李凯周宇荀
Owner DONGHUA UNIV
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