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Anti-bacterial peptide PD22

A PD22, antibacterial peptide technology, applied in multiple fields, can solve the problems of limiting the popularization and application of antibacterial peptides, host cytotoxic side effects, low antibacterial activity, etc., and achieve the effects of convenient artificial synthesis, small molecular weight and low cytotoxicity

Active Publication Date: 2015-11-18
FUJIAN AONONG BIOLOGICAL TECH GRP CO LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, most of the antimicrobial peptides from natural sources have defects such as large molecular weight, low antibacterial activity, toxic side effects on host cells, or hemolysis of the host, which limits the popularization and application of antimicrobial peptides in weaned piglets to a certain extent.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Embodiment 1: Preparation of antimicrobial peptide PD22

[0016] The chemical synthesis method of the antimicrobial peptide PD22: according to the amino acid sequence described in the summary of the invention, the full sequence of PD22 (synthesized by the School of Life Sciences, Hunan Normal University) was synthesized with an automatic peptide synthesizer (ABI433), and purified by desalting and desalting by HPLC reverse-phase column chromatography ;

[0017] The molecular weight of the purified antimicrobial peptide PD22 was determined by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF), the isoelectric point was determined by isoelectric focusing electrophoresis, and the amino acid sequence was analyzed by an automatic amino acid sequencer.

[0018] Antimicrobial peptide PD22, the antimicrobial peptide PD22 is an artificially designed and synthesized active polypeptide, comprising 13 amino acid residues, a molecular weight of 1...

Embodiment 2

[0019] Embodiment 2: antimicrobial peptide PD22 antibacterial experiment

[0020] The antibacterial effect was reflected by measuring the minimum inhibitory concentration (MIC) of the antimicrobial peptide PD22, which was the lowest sample concentration at which no bacterial growth could be detected. Using the double dilution method, the specific method is as follows:

[0021] The test bacteria were inoculated on LB solid medium, and then cultured upside down in a 37°C incubator. After the colonies grow out, use a disposable inoculation loop to pick a single colony of each test bacterium into LB liquid medium, and culture it in a 37°C incubator with shaking until the logarithmic growth phase. Detect the OD600 of the bacterial solution on the ultraviolet spectrophotometer, according to 1OD600=1×10 9 CFU / mL Dilute the test bacteria liquid with liquid LB medium to 2×10 5 CFU / mL. Then add 100 μ L LB liquid medium to each well of a sterile 96-well plate, and then add 100 μ L of...

Embodiment 3

[0026] Embodiment 3: Cytotoxicity experiment of antimicrobial peptide PD22

[0027] Pig small intestinal epithelial cells IPEC-1 (provided by the Institute of Subtropical Agroecology, Chinese Academy of Sciences) were cultured in a medium containing 5% Tai bovine serum, 200U / mL double antibody (penicillin and streptomycin each 100U / mL), 5ug / L epidermal growth factor ( EGF) cultured in HAM'S / F12 medium to logarithmic phase, washed three times with phosphate buffer (HyClone), added 0.25% (2.5g / L) trypsin to digest to a single cell, added HAM'S / F12 culture Centrifuge at 400g for 4 minutes after the base, discard the supernatant, suspend the cells with fresh HAM'S / F12 medium, and adjust the cell density to 1×10 6 cells / mL, then add 200uL to each well of a 96-well cell culture plate, add different concentrations of antimicrobial peptide PD22 after the cells adhere to the wall, and place in a carbon dioxide incubator (37°C, 5% CO 2 ) after culturing for 24 hours, add 20uLCCK-8 (Bei...

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Abstract

The invention discloses anti-bacterial peptide PD22. The amino acid sequence of the anti-bacterial peptide PD22 is shown in SEQ ID NO:1. The anti-bacterial peptide PD22 has the advantages of being small in molecular weight, capable of being conveniently and artificially synthesized, low in cost, strong in killing effect on escherichia coli, salmonella, clostridium welchii and other bacteria, low in clostridium welchii and capable of remarkably decreasing the diarrhea rate of weaned pigs and increasing the growth speed of weaned pigs after being applied to weaned pig feed.

Description

technical field [0001] The invention relates to multiple fields, in particular to an antimicrobial peptide PD22. Background technique [0002] In order to improve the reproductive efficiency of sows and reduce the vertical transmission of maternal diseases, piglets in pig farms in my country generally choose to be weaned at 3-4 weeks of age, while the weaning process generally occurs at 3-4 months in nature. Early weaning of piglets caused great stress to piglets. Piglets were separated from sows, nutritional sources changed from liquid breast milk to solid feed, mixed groups and environmental changes, and the digestive system and immune system of early weaned piglets were not fully developed. It is very easy to be attacked by various pathogenic bacteria and infected with diseases, among which digestive tract disease is one of the most common and serious diseases, and it is also an important cause of growth inhibition and even death of weaned piglets. For a long time, the u...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/08C07K1/20A23K1/16A23K1/18
Inventor 杨焕胜周盛昌肖丽萍施建成杨林姜文叶俊标
Owner FUJIAN AONONG BIOLOGICAL TECH GRP CO LTD
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