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Method for screening feed antibiotic substitute product and product thereof

A technology for substitutes and antibiotics, applied in biochemical equipment and methods, material inspection products, chemical instruments and methods, etc., can solve the problems of high cost, long screening and identification cycle, low efficiency, etc., to improve production performance, inhibit Intestinal diarrhea, the effect of improving vegetative growth

Active Publication Date: 2018-11-02
HUNAN AGRICULTURAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, my country's aquaculture and feed industries are actively preparing to deal with the "post-antibiotic" era, in order to reduce the impact of the restriction or banning of feed antibiotics on farming in the future, and finding alternatives to feed antibiotics is the most important solution, but Due to the wide variety of potential feed antibiotic substitutes, the screening and identification through animal experiments has a long cycle, low efficiency, and high cost. Therefore, it is particularly important to establish an efficient, fast, and low-cost screening method and technology platform.

Method used

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  • Method for screening feed antibiotic substitute product and product thereof
  • Method for screening feed antibiotic substitute product and product thereof
  • Method for screening feed antibiotic substitute product and product thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 2

[0086] Example 2. Assessment of the Antibiotic Potential of Alpha-Ketoglutarate

[0087] The stably transformed PXR porcine intestinal epithelial cell line pxr_IPEC-J2 obtained by the method described in the patent was used as the research object to evaluate the substitution potential of alpha-ketoglutarate. The specific implementation is as follows: 1) The stable transfection cell line pxr_IPEC-J2 is cultured in the whole medium, and the cells are cultured to the logarithmic phase (80-90% fusion) for subculture and plating. 2) After the cells grow to the logarithmic phase (80-90% confluence), the experimental group is treated with alpha-ketoglutarate at different concentrations (1.0 mM, 2.0 mM, 5.0 mM) for 48 hours. 3) Collecting cells, extracting mRNA and total cell protein respectively. 4) Real time PCR technology was used to detect the expression of pgPXRmRNA in porcine intestinal cell lines. Second, the expression of pgPXR protein was detected by western blot technique....

Embodiment 3

[0088] Example 3. Assessment of the Antibiotic Potential of Alliin (Precursor of Allicin)

[0089] The stably transformed PXR porcine intestinal epithelial cell line pxr_IPEC-J2 obtained by the method described in the patent was used as the research object to evaluate the alliinin substitution potential. The specific implementation is as follows: 1) The stable transfection cell line pxr_IPEC-J2 is cultured in the whole medium, and the cells are cultured to the logarithmic phase (80-90% fusion) for subculture and plating. 2) When the cells grow to the logarithmic phase (80-90% fusion), the experimental group is added with different concentration gradients (1.0ug / mL, 2.0ug / mL, 5.0ug / mL,) of alliin, and the control group is added with the experimental The same amount of DMSO was treated for 48 hours. 3) Collecting cells, extracting mRNA and total cell protein respectively. 4) Real time PCR technology was used to detect the expression of pgPXR mRNA in porcine intestinal cell lin...

Embodiment 4

[0090] Embodiment 4, the potential evaluation of the replacement potential of Boluohui

[0091] The stably transformed PXR porcine intestinal epithelial cell line pxr_IPEC-J2 obtained by the method described in the patent was used as the research object to evaluate the potential of replacing Boluohui. The specific implementation is as follows: 1) The stable transfection cell line pxr_IPEC-J2 is cultured in the whole medium, and the cells are cultured to the logarithmic phase (80-90% fusion) for subculture and plating. 2) When the cells grow to the logarithmic phase (80-90% confluence), the experimental group is added with different concentration gradients (1.0ug / mL, 2.0ug / mL, 5.0ug / mL,) and the control group is added to the experimental group. The same amount of DMSO was treated for 48 hours. 3) Collecting cells, extracting mRNA and total cell protein respectively. 4) Real time PCR technology was used to detect the expression of pgPXR mRNA in porcine intestinal cell lines. ...

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Abstract

The invention belongs to the technical field of genetic recombination, and concretely discloses a method for screening an antibiotic substitute product by building a pig intestinal tract PXR (pgPXR) cell platform through recombinant plasmids. The process comprises the following steps of target gene pgPXR extraction, amplification, purification recovery, digestion, connection, conversion and identification. The material used in the method mainly comprises a pig jejunum endothelial cell line IPEC-J2 and pCI-neo connection carrier and DH5a competent cells. By using the technology, the pig intestinal tract cell line for stably expressing PXR receptors can be obtained; the platform can be used as an effective antibiotic substitute product screening platform.

Description

technical field [0001] The invention relates to the field of gene recombination technology and the field of drug screening, more specifically, to a method for constructing and linking expression of PXR plasmids and a method for drug screening based on intestinal PXR signal activation. Background technique [0002] Pregnane X receptor (PXR) is a member of the nuclear receptor superfamily (NR), and its main biological function is to participate in the body's metabolism and excretion of endogenous and exogenous compounds, thereby maintaining the body's The homeostasis of the internal environment has been widely concerned in the medical field because it has been found to have important biological and pathological functions in recent years. Previous studies have shown that the main biological effect of PXR is the "detoxification" function, that is, the removal of heterologous and endogenous toxic substances, which is an important part of maintaining the homeostasis of the body. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N15/85C12Q1/686G01N33/68
CPCC07K14/70567C12N15/85C12Q1/686G01N33/68C12Q2561/113
Inventor 符晨星贺建华方俊伍小松姚康
Owner HUNAN AGRICULTURAL UNIV