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Kit for detecting sheep border diseases

A border disease and kit technology, applied in the field of kits for detecting sheep border disease, can solve the problem of inability to detect sheep border disease early and achieve rapid and effective detection

Inactive Publication Date: 2018-11-23
刘欢
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Therefore, if the sheep border disease can be diagnosed in time in the early stage of the onset, the treatment effect will be greatly improved, and the existing routine detection methods cannot effectively detect the sheep border disease in the early stage. Therefore, it is urgent to provide an early diagnosis of sheep border disease. borderline disease approach

Method used

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  • Kit for detecting sheep border diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0013] Embodiment 1 sample collection

[0014] Blood was collected from 50 sheep with borderline disease and 25 normal sheep.

Embodiment 2

[0015] The extraction of embodiment 2 sample total RNA

[0016] 1. Extraction of serum total RNA

[0017] 1) Add 3 times the volume of TRIzol (0.75ml) to 0.25ml of the collected blood sample, shake and mix well.

[0018] 2) Place the homogenized sample at 15-30°C for 5 minutes.

[0019] 3) Centrifuge for 10 min (12000 rpm, 4° C.), and take the supernatant.

[0020] 4) Add 0.2ml of chloroform, cover the tube cap, shake vigorously for 15 seconds, and place at room temperature for 3 minutes.

[0021] 5) Centrifuge for 15 minutes (12000 rpm, 4°C). Transfer the upper aqueous phase (about 500 μl) to a new tube.

[0022] 6) Add an equal volume of isopropanol to the obtained aqueous phase solution, mix well, and place at room temperature for 30 minutes.

[0023] 7) Centrifuge for 10 minutes (12000 rpm, 4° C.), and remove the supernatant. After centrifugation, RNA forms a gel-like pellet on the side and bottom of the tube.

[0024] 8) Add 1ml of 75% ethanol to wash the precipita...

Embodiment 3

[0044] Example 3 Fluorescent quantitative PCR detection of BDV expression in sheep border disease and control group

[0045] 1) Fluorescence quantitative PCR

[0046] The kit uses FastFire qPCR PreMix (SYBR Green) from Tiangen Company. First, thaw FastFire qPCR PreMix, templates, primers, and RNase-free water, and dissolve all reagents at room temperature and mix thoroughly. Next, a fluorescent quantitative PCR system was prepared.

[0047] Fluorescent quantitative PCR system

[0048] Composition

concentration

Volume (μl)

2*FastFire qPCR PreMix

1*

10

Forward primer (10μM)

300nM

0.6

Reverse primer (10μM)

300nM

0.6

cDNA template

-

1

RNase-free water

-

7.8

total capacity

-

20

[0049] For BDV expression detection, three parallel reactions were set up for each sample, and snRNA U6 was used as an internal reference.

[0050] The forward primer sequence in the above table is: AC...

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Abstract

The invention relates to a kit for detecting sheep border diseases. The kit comprises a reagent for specifically detecting a sheep border disease virus BDV. The kit provided by the invention can be used for effectively detecting the sheep border disease virus; the kit can be used for realizing quantitative detection of the BDV, so that the aim of rapidly and effectively detecting the sheep borderdisease virus is realized.

Description

technical field [0001] The invention relates to the field of veterinary medical detection, in particular to a kit for detecting sheep border disease. Background technique [0002] Border disease (BD), named after it was first discovered in the border area between England and Wales, also known as Hairyshaker disease of lambs, is caused by border disease virus (BDV) A congenital infectious disease of newborn lambs characterized by hirsutism, poor growth and neurological abnormalities. Sheep border disease virus has immunosuppressive effect. After border disease virus infection during pregnancy, the virus can persist in various tissues in the body and become the carrier of the virus and the source of disease regardless of whether the fetus has immune response ability. Infected lambs are still infective to their offspring within a few years after they grow to maturity. The virus present in the germ cells of the uterus and ovaries or testes can be vertically transmitted through ...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6851C12R1/93
CPCC12Q1/701C12Q1/6851C12Q2531/113C12Q2563/107C12Q2521/107
Inventor 刘欢
Owner 刘欢
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