Unlock instant, AI-driven research and patent intelligence for your innovation.

cahv-tnfr specific gene and its application

A specific gene and gene technology, applied in the fields of fishery biotechnology, aquatic animal medicine, and virology, can solve problems such as difficult identification and diagnosis, achieve efficient and accurate diagnosis, reduce costs, and improve efficiency

Active Publication Date: 2019-03-19
INST OF AQUATIC LIFE ACAD SINICA
View PDF10 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in areas where both CaHV and CyHVs are prevalent, it is difficult to differentiate and diagnose

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • cahv-tnfr specific gene and its application
  • cahv-tnfr specific gene and its application
  • cahv-tnfr specific gene and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] A diagnostic kit for gill hemorrhage of crucian carp, including primers as shown in Table 1:

[0043] Table 1 Basic information of primers

[0044]

[0045] The above primers are based on the comparison of the genome and homologous gene sequences of the crucian herpes virus CaHV (Carassius aruatus herpesvirus, the present invention may be called acute crucian gill hemorrhagic disease herpes virus) and carp herpes virus CyHVs, and are selected for the main capsid protein CaHV-MCP of CaHV A pair of primers CaHV-MCP-F / R and CaHV-TNFR-F / R were designed for the high identity sequence of gene 88R and the low identity sequence of CaHV tumor necrosis factor CaHV-TNFR gene 146R. That is, according to the corresponding sequence of Ca HV-MCP gene 88R ( figure 1 ), designed and synthesized a pair of primers CaHV-MCP F and CaHV-MCP R (this pair of primers is abbreviated as CaHV-MCP F / R); meanwhile, according to the corresponding sequence of CaHV-TNFR gene 146R ( figure 2 ), de...

Embodiment 2

[0048] A kind of application of crucian carp gill hemorrhage diagnosis kit, application process comprises:

[0049] 1) Extract the DNA of the sample to be tested:

[0050] Using a DNA extraction kit (TaKaRa MiniBEST Universal Genomic DNA Extraction KitVer.5.0, product of Clontech Company), the DNA of the sample to be tested was extracted as template DNA for multiplex PCR amplification. For DNA extraction steps, please refer to the kit instructions.

[0051] 2) Multiplex PCR amplification and product analysis

[0052] First add various reaction components into the PCR reaction tube, then set the cycle parameters, and then carry out cycle amplification. The specific method is as follows: add 1 μl (0.1-1.0 μg) of the sample DNA to be tested in step 1) to the PCR reaction tube, add 0.5 μl 10 μM each of the primers CaHV-M CPF / R and CaHV-TNFR F / R; add 1 μl , 10mM dNTP Mix; add 2.5μl 1 0×TranStart Taq Buffer (Mg 2+ ) and 0.5 μl TranStart Taq DNA Polymerase (product of Beijing Qua...

Embodiment 3

[0058] Specificity of diagnostic kit for gill hemorrhage in crucian carp

[0059] 1) Template nucleic acid sample preparation:

[0060] Extraction of non-herpesvirus genomic nucleic acid from different aquatic animals: according to the chloroform-phenol extraction method [Chen et al. Veterinary Research 2013,44:101], the genomic RNA of mandarin fish rhabdovirus (Siniperca chuatsirhabdovirus, SCRV) and swamp green bullfrog were respectively prepared Rana grylio virus (RGV) genomic DNA, and giant salamander frog virus (Andrias davidianus ranavirus, ADRV) genomic DNA. Genomic RNA of fish rhabdovirus was amplified by reverse transcription by RT-PCR, and cDNA was synthesized from RNA. Then, the cDNA, RGV genome DNA and ADRV genome DNA synthesized by reverse transcription of the SCRV genome were respectively used as templates of non-herpes virus genomes of aquatic animals, and served as negative controls.

[0061] Genomic DNA of acute carassius aruatus herpesvirus (Carassius aruat...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a CaHV-TNFR (carassius aruatus herpesvirus-tumor necrosis factor receptor) specific gene and an application. The CaHV-TNFR specific gene sequence is represented as SEQ ID NO.1. Primers for carassius auratus gill hemorrhage disease is designed according to the gene and include CaHV-MCP F / R and CaHV-TNFR F / R. The two pairs of primers are adopted to perform multiplex PCR (polymerase chain reaction) amplification, and when two fragments of 436bp and 915bp are amplified simultaneously, the condition that a sample is infected with CaHV can be diagnosed; when only one fragment of 436bp is amplified, the condition that the sample is infected with CyHVs (Cyprinid herpesviruses) can be detected. The gene sequence and the primers are particularly suitable for genetic diagnosis and molecular epidemiological investigation of acute carassius auratus gill hemorrhage herpesvirus disease and are applicable to monitoring and early warning of herpesviruses of aquatic animals, the positive detection rate and the accuracy rate are both higher than 98%, and the herpesviruses can be detected when CaHV gene quantity of the to-be-detected sample is 10 pg / mu L.

Description

technical field [0001] The invention belongs to the fields of virology, fishery biotechnology and aquatic animal medicine, and more specifically relates to a CaHV-TNF R specific gene, and also relates to the use of a CaHV-TNFR specific gene. Background technique [0002] The World Organization for Animal Health (OIE) has listed fish herpes virus as a list of key aquatic animal diseases, and my country has also included it in the list of second-class diseases [Administrative Administration of Fisheries, Ministry of Agriculture. 2014 China Aquatic Animal Health Status Report. China Agricultural Press, Beijing, 2015]. Herpesviruses are a class of enveloped viruses with large double-stranded linear DNA genomes. It is known that the genus Cyprinivirus is a member of the family Alloherpesviridae, known as Cyprinid herpesviruses, including types 1, 2 and 3 (CyHV1, CyHV2 and CyHV3)[ Zhang Qiya, Gui Jianfang. Chinese Science: Life Science, 2014, 44:1236–1252], which can be abbreviat...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/38C12N15/11C12Q1/70C12Q1/686C07K16/08G01N33/577G01N33/569C12R1/93
CPCC07K14/005C07K16/085C12N2710/16022C12Q1/705C12Q2600/158C12Q2600/16
Inventor 张奇亚曾小涛陈中元
Owner INST OF AQUATIC LIFE ACAD SINICA