Animal response type gene under microbial infection, identification and applications thereof

A microbial infection and microbial technology, applied in the biological field, can solve the problems of body disease, difficulty in achieving effective sterilization, and difficulty in transporting antibiotics

Inactive Publication Date: 2015-07-22
JIANGSU MICROMEDMARK BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The main reasons include: when these pathogens invade and lurk in the cells, it is difficult for conventional antibiotics to transport to the cells where the bacteria are dormant, and even if they are transported, the concentration is difficult to achieve an effective bactericidal effect; antibodies canno...

Method used

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  • Animal response type gene under microbial infection, identification and applications thereof
  • Animal response type gene under microbial infection, identification and applications thereof
  • Animal response type gene under microbial infection, identification and applications thereof

Examples

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preparation example Construction

[0109] The preparation of the miRNA chip can adopt conventional manufacturing methods of biochips known in the art. For example, if the solid phase carrier is a modified glass slide or silicon wafer, and the 5' end of the probe contains amino-modified poly dT strings, the oligonucleotide probe can be formulated into a solution, and then spotted with a spotting instrument. The miRNA chip of the present invention can be obtained by arranging in a predetermined sequence or array on a modified glass slide or a silicon chip, and then fixing it by standing overnight.

[0110] The solid-phase hybridization between the RNA of the present invention and the miRNA chip is carried out according to the classic method in this field, and those skilled in the art can easily determine relevant buffer solution, probe and sample concentration, pre-hybridization temperature, hybridization temperature and time etc. based on experience. the optimum conditions. Or you can also refer to the "Molecul...

Embodiment 1

[0138] Identification and application of miRNAs in response to Mycobacterium tuberculosis

[0139] (1) Identification of response miRNAs in Mycobacterium tuberculosis-infected cells

[0140] 1.1 Cell culture

[0141] In antibiotic-free RPMI medium, use a 24-well plate to infect dendritic cells of C57BL / 6 mice (from the Cell Bank of the Chinese Academy of Sciences) with Mycobacterium tuberculosis (from the School of Life Sciences, Nanjing University) at a ratio of 1:10 (DC); at 5% CO 2 In the incubator, cultivate at 37°C. Dendritic cells without any treatment (U), treated with 1 μg / ml sugar polylipid (LPS), infected with H37Rv and treated with LPS (LPS+H37RV) were used as controls.

[0142] 1.2 Screening of response miRNAs in cells infected with Mycobacterium tuberculosis

[0143] Firstly, the total RNA was extracted using the miRNA mini-extraction kit (purchased from Shanghai Yubo Biotechnology Co., Ltd.), and then the samples were detected by EXIQON (purchased from Vedbae...

Embodiment 2

[0161] Screening and Identification of H1N1 Responsive miRNAs

[0162] In this example, the research on influenza virus H1N1 (obtained from the School of Life Sciences of Nanjing University) showed that after H1N1 infection, the level of miRNA in animal cells would change in response. And the use of these responsive miRNAs can assist in the diagnosis of infectious agents / has a high diagnostic value.

[0163] (1) Screening of response miRNAs in virus-infected cells

[0164] Blood samples were centrifuged using a Ficoll density gradient centrifuge to obtain human peripheral blood mononuclear cells (PBMC), which were stored in RNA sample preservation solution RNAlater (Ambion) at -80°C for RNA extraction; then the miRNA mini-extraction kit ( Total RNA was extracted from peripheral blood mononuclear cells (purchased from Shanghai Yubo Biotechnology Co., Ltd.).

[0165] Infected samples and healthy controls were tested using a biochip method. The specific operation is as follows...

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Abstract

The present invention relates to an animal response type gene under microbial infection, identification and applications thereof, and particularly provides a method for non-diagnostic identification of animal body miRNA. The method comprises: (a) providing a test group and a control group, wherein the test group is the animal individual, the animal sample or the animal cells being subjected to any one microbial infection; (b) comparing the expression profiles of the miRNA derived from the animals in the test group and the control group so as to identify the response type miRNA; (c) comparing the identified response type miRNA and the microbial gene database so as to determine the type of the response type miRNA having the target relationship with the microbial; and (d) verifying the regulating relation on the type of the response type miRNA having the target relationship through experiments.

Description

technical field [0001] The invention relates to the field of biotechnology, and more specifically relates to response genes of animals under microbial infection, identification and application thereof. Background technique [0002] Microorganism is a general term for a group of tiny organisms widely present in nature that are invisible to the naked eye and must be magnified hundreds, thousands or even tens of thousands of times with the help of an optical microscope or an electron microscope. They have the characteristics of small size, simple structure, rapid reproduction, easy variation and strong adaptability to the environment. [0003] There are many kinds of microorganisms, which are widely distributed in nature. There are also a variety of microorganisms on the body surface of humans, animals and plants and in the cavities that communicate with the outside world. [0004] Microorganisms mainly include: bacteria, archaea, protists, viruses, etc. [0005] One of the ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/113A61K48/00A61K45/00A61P31/00
Inventor 张辰宇曾科王进陈熹
Owner JIANGSU MICROMEDMARK BIOTECH
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