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A kind of ssc-mir-122-5p associated with c-type Clostridium perfringens infectious diarrhea in piglets and its application

A technology for Clostridium perfringens and infectious diarrhea, applied in ssc-miR-122-5p and its application fields, can solve the problem of less research on the mechanism of action

Active Publication Date: 2022-06-21
GANSU AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there are few studies on the mechanism of action of miRNA in bacterial infectious diarrhea in piglets.

Method used

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  • A kind of ssc-mir-122-5p associated with c-type Clostridium perfringens infectious diarrhea in piglets and its application
  • A kind of ssc-mir-122-5p associated with c-type Clostridium perfringens infectious diarrhea in piglets and its application
  • A kind of ssc-mir-122-5p associated with c-type Clostridium perfringens infectious diarrhea in piglets and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Embodiment 1, the expression detection of ssc-miR-122-5p

[0022] 1. Detection of the expression of ssc-miR-122-5p in the ileum tissue of piglets with Clostridium perfringens type C infectious diarrhea

[0023] Step 1. Sample collection

[0024] 30 healthy 7-day-old piglets (long × big) were selected, and 5 piglets were randomly selected as the control piglet group (IC), fed with the same dose of culture medium without the inoculated strain, and the remaining 25 piglets were treated with C-type perfringens Bacteria oral challenge test, each piglet was fed with C-type Clostridium perfringens C59-2 strain culture solution (1×10 9 CFU / mL) 1 mL, and the test period was 5 d. Immediately collect ileal tissue samples and store them in liquid nitrogen.

[0025] Step 2. Total RNA extraction from ileal tissue

[0026] Take out the tissue samples from liquid nitrogen, grind them into powder, collect them in a 1.5 mL centrifuge tube, add 1 mL of TransZol Up, shake and mix well,...

Embodiment 2

[0046] Example 2. Effect of overexpression and interference of ssc-miR-122-5p on inflammatory injury of IPEC-J2 cells treated with CPB2 toxin

[0047] 1. Overexpression of ssc-miR-122-5p and detection of interference efficiency

[0048] Step 1. IPEC-J2 cell culture

[0049] The steps are the same as in Example 1.

[0050] Step 2. Chemical synthesis of miRNA mimics and inhibitors

[0051] Based on the sequence information of porcine ssc-miR-122-5p provided by the miRBase database, miRNA mimics and inhibitors were chemically synthesized in vitro. The synthesized small RNA fragments were designed and synthesized by Shanghai Gema Pharmaceutical Technology Co., Ltd. The synthetic sequence of ssc-miR-122-5pmimic is shown as SEQ ID NO.3-SEQ ID NO.4, and the synthetic sequence of ssc-miR-122-5p inhibitor is shown as SEQ ID NO.5.

[0052] Step 3. Cell Transfection

[0053] IPEC-J2 cells were seeded in 6-well culture plates, and when the cell density reached 70%, washed with PBS fo...

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Abstract

The invention discloses a ssc-miR-122-5p related to piglet type C Clostridium perfringens infectious diarrhea, the nucleotide sequence of the ssc-miR-122-5p is shown in SEQ ID NO.1 . The present invention also discloses specific primers for detecting the expression level of ssc-miR-122-5p, the sequence of which is shown in SEQ ID NO.2. The invention discloses a method for improving piglet resistance to type C Clostridium perfringens infection by interfering with ssc-miR-122-5p. The present invention found for the first time that ssc-miR-122-5p was significantly highly expressed in the ileum tissue of piglets with Clostridium perfringens type C infectious diarrhea, and then IPEC-J2 cells were used as the research object to overexpress and interfere with ssc-miR ‑122‑5p studied its effect on CPB2 toxin-induced IPEC‑J2 cell damage, in order to elucidate the mechanism of ssc‑miR‑122‑5p involved in the regulation of diarrhea caused by Clostridium perfringens type C infection, which is a bacterium in piglets Molecular breeding of sexual diarrhea provides a powerful scientific reference.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to ssc-miR-122-5p related to piglet C-type Clostridium perfringens infectious diarrhea and its application. Background technique [0002] At present, Clostridium perfringens type C has become the main cause of intestinal infection of various livestock and poultry in modern breeding industry, especially for piglets within one week of age, it can cause hemorrhagic diarrhea, and then bacteria and toxins enter the body circulatory system, leading to multiple organ failure and ultimately death of the body. According to statistics, the morbidity and mortality of piglet diarrhea caused by Clostridium perfringens type C can be as high as 100%. At present, the incidence of piglet diarrhea has been reduced by means of antibiotics, vaccines, etc., and with the overuse of antibiotics, not only the structure of the intestinal mucosa of piglets is changed, the intestinal barrier is damag...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883C12N15/113C12N15/11A61K31/7088A61P1/12A61P31/04
CPCC12Q1/6883A61K31/7088A61P1/12A61P31/04C12Q2600/158C12Q2600/178
Inventor 张生伟滚双宝高小莉杨巧丽
Owner GANSU AGRI UNIV
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