Biliverdin preparation and application thereof in prevention and treatment of porcine reproductive and respiratory syndrome, and detection method

A technology for respiratory syndrome and biliverdin, which is applied in the field of biomedicine to achieve obvious effects, convenient use, and easy transportation

Active Publication Date: 2017-03-08
NORTHWEST A & F UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The present invention seeks a kind of medicine that has good antiviral effect to PRRSV in order to overcome the problem ...

Method used

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  • Biliverdin preparation and application thereof in prevention and treatment of porcine reproductive and respiratory syndrome, and detection method
  • Biliverdin preparation and application thereof in prevention and treatment of porcine reproductive and respiratory syndrome, and detection method
  • Biliverdin preparation and application thereof in prevention and treatment of porcine reproductive and respiratory syndrome, and detection method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] The preparation of embodiment 1 biliverdin

[0034] Dissolve biliverdin (purchased from Sigma, USA) in 0.1 mol / L NaOH, adjust the pH value to 7.4 with 1 mol / L HCL, then dilute with PBS, filter and sterilize with a 0.2 μM filter membrane, and separate Wrap it with tinfoil and store at -80°C for later use.

Embodiment 2

[0035] Example 2 Biliverdin Treatment of Marc-145 Cells Infected by PRRSV

[0036] Marc-145 cells at 1×10 5 The density of cells / ml was cultured in a six-well cell culture plate containing DMEM medium (containing penicillin 100 U / ml, streptomycin 50 μg / ml, 10% fetal bovine serum), 37 ° C, 5% CO 2 Culture in a humid incubator until 70%-80% confluent, discard the medium. Insert 0.1 MOI of PRRSV into each well, incubate at 37°C for 1 h, discard unadsorbed virus liquid, and wash 3 times with 1 ml PBS / well. Add 2ml of culture medium containing different concentrations (10 μM, 20 μM, 50 μM, 100 μM, 150 μM) of biliverdin diluted in Example 1 to each well, place at 37° C., 5% CO 2 cultured in a humidified incubator. 48h after virus infection, the cell supernatant and cells were collected respectively.

Embodiment 3

[0037] Example 3 qPCR detection of PRRSV ORF7 mRNA expression after treatment with different concentrations of biliverdin

[0038] After the cells treated in Example 2 were digested with trypsin, they were collected in a 1.5 ml centrifuge tube, centrifuged at 500 g at 4° C. for 10 min, and the supernatant was discarded. 1ml TRIzol (purchased from Invitrogen) was added to each tube, and total RNA was extracted according to the instructions. RNA was reverse-transcribed into cDNA using PrimeScript RT reagent Kit Perfect Real Time kit (purchased from TAKARA).

[0039] Finally, the SYBR GREEN kit (purchased from Roche Company) and the above reverse-transcribed cDNA template and the designed upstream and downstream primers of the PRRSV ORF7 gene were used to perform real-time fluorescent quantitative PCR detection on the StepOnePlus Real-Time PCRSystem of Applied Biosystems.

[0040] The reaction conditions were as follows: denaturation at 95°C for 10 min; 40 cycles of reaction at ...

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Abstract

The invention discloses a biliverdin preparation and an application thereof in prevention and treatment of PRRSV, and a detection method. An antiviral effect of exogenous biliverdin on the PRRSV is researched through qRT-PCR, western blot, cell supernatant TCID50 detection and indirect immunofluorescence assay and other methods in Marc-145 and PAM cell levels. Results show that biliverdin significantly inhibits proliferation of the PRRSV in susceptible cells and host target cells. The biliverdin preparation disclosed by the invention has good antiviral effect, and is expected to become a novel drug for prevention and control of porcine reproductive and respiratory syndrome.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and the invention relates to a biliverdin preparation and its application and detection method, in particular to a biliverdin preparation and its application and detection method in anti-PRRSV. Background technique [0002] Porcine reproductive and respiratory syndrome (PRRS), also known as porcine blue ear disease, is caused by porcine reproductive and respiratory syndrome virus (Porcine reproductive and respiratory syndrome virus, PRRSV), which is extremely harmful to the global pig industry. important infectious diseases of pigs. PRRS mainly causes abortion, stillbirth, mummified fetus, weak piglets, and respiratory symptoms in pigs of all ages, especially piglets. The characteristic lesion is interstitial pneumonia and the mortality rate is extremely high. [0003] The prevention and control of PRRSV is a difficult problem in my country and the world at present. PRRSV is difficult to pr...

Claims

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Application Information

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IPC IPC(8): A61K31/4025A61K9/19A61P31/14G01N33/50
CPCA61K9/0019A61K9/19A61K31/4025G01N33/5055
Inventor 肖书奇周恩民张昂克
Owner NORTHWEST A & F UNIV
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