Gene chip for detecting peste des petits ruminants virus and detecting method for gene chip

A technology of Peste des petits ruminants and gene chips, which is applied in the direction of microorganism-based methods, biochemical equipment and methods, and the determination/inspection of microorganisms, can solve the problems of lack of detection methods for PPR virus gene chips, and achieve stability and economical efficiency. Time-saving, labor-saving, strong stability and high sensitivity

Inactive Publication Date: 2015-05-13
INSPECTION & QUARANTINE TECH CENT OF CHONGQING ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, gene chip detection technology has been applied to the detection of many diseases, but there is a lack of detection methods for the gene chip of Peste des petits ruminants virus

Method used

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  • Gene chip for detecting peste des petits ruminants virus and detecting method for gene chip
  • Gene chip for detecting peste des petits ruminants virus and detecting method for gene chip
  • Gene chip for detecting peste des petits ruminants virus and detecting method for gene chip

Examples

Experimental program
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Effect test

Embodiment 1

[0036]Example 1, the present invention is based on the cell proliferation of Peste des petits ruminants virus, and the virus genome is extracted. Select different specific conserved sequences to design an oligonucleotide probe, which can specifically bind to the PCR product of the corresponding viral cDNA. When performing genome amplification by PCR, the 5′ end of the universal upstream primer is labeled with Cy3 fluorescent dye , the probe can hybridize with this product at 50°C. The hybridization result is judged according to the position and intensity of the fluorescent signal, so as to detect Peste des petits ruminants virus. PCR primers SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6 and SEQ ID NO.7 were designed using Primer Premier 5.0 biological software according to the gene sequence of Peste des petits ruminants virus published on the Genbank website. The gene sequence was cloned, sequenced, compared and analyzed, and the conserved sequence was selected to design a specific o...

Embodiment 2

[0044] Example 2, see figure 1 , the gene chip used for the detection of Peste des petits ruminants virus, adopts the micro-spotting technology of the gene chip, immobilizes the probes of the samples to be tested and various quality control probes on the chemically modified aldehyde-based substrate, forming 7 rows × 8 columns microarray. The distribution of probes on the microarray from top to bottom is as follows: spotting position quality control partition P1: 1 row × 8 points, hybridization positive quality control partition P2: 1 row × 4 points, foot-and-mouth disease virus type A detection probe partition 1: 1 line × 4 points, FMD virus Asian type detection probe division 2: 1 line × 4 points, FMD virus general detection probe division 3: 1 line × 4 points, FMD virus O-type detection probe division 4: 1 line × 4 points, vesicular stomatitis virus detection probe division 5: 1 row × 4 points, porcine vesicular disease virus detection probe division 6: 1 row × 4 points, Pe...

Embodiment 3

[0047] Embodiment 3, reagent preparation

[0048] 1) Chip washing solution

[0049] According to needs and actual conditions, prepare washing solution I and washing solution II according to the following proportions.

[0050] Washing solution Ⅰ: the final concentration of SSC is 2×, and the final concentration of SDS is 0.2%. For example, 500mL lotion I = 440mL distilled water + 50mL 20×SSC + 10mL 10% SDS, or prepare in proportion as required.

[0051] Washing solution Ⅱ: The final concentration of SSC is 0.2×. Such as 500mL lotion II = 495mL distilled water + 5mL 20 × SSC, or according to the need to prepare in proportion.

[0052] If 10% SDS produces white flocculent precipitates, please dissolve and mix in a 42°C water bath to prepare a lotion.

[0053] 2) absolute ethanol.

[0054] 3) Ice-water mixture.

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Abstract

The invention discloses a gene chip for detecting a peste des petits ruminants virus and a detecting method for the gene chip. The gene chip comprises a PCR (polymerase chain reaction) mixed liquor tube, Taq polymerase, PCR positive control, PCR negative control, ddH2O, a detecting chip, point sample distribution of a chip probe, a chip cover plate, a chip rail, a hybridization solution, a hybrid positive control and a hybrid box. A PCR primer is analyzed and designed by a standard strain genomic sequence to perform target gene cloning and sequencing analysis, and then the specific probe is designed to detect the peste des petits ruminants virus. The invention aims to a micro array chip which is high in sensitivity, strong in specificity, time-saving, labor-saving and easy to observe results for detecting the peste des petits ruminants virus.

Description

technical field [0001] The invention belongs to the field of biological chips. Specifically, the present invention relates to a gene chip of Peste des petits ruminants virus and a detection method thereof. Background technique [0002] Peste des petits ruminants (PPR) is an acute, febrile and highly contagious infectious disease of small ruminants such as goats and sheep caused by peste des petits ruminants virus (PPRV). sheep plague" or "pseudo-rinderpest". The main clinical symptoms of the disease are high fever retention, increased eye and nose secretions, oral erosion, diarrhea and pneumonia. The infection rate of susceptible animals is 100%, and the mortality rate can reach 50%-100%. The World Organization for Animal Health (OIE) lists it as a class A animal disease, and the "List of Entering Animal Quarantine Diseases of the People's Republic of China" (Announcement No. 2013) jointly issued by the Ministry of Agriculture and the General Administration of Quality Supe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
CPCC12Q1/70C12Q1/6837C12Q2565/501C12Q2531/113
Inventor 王昱聂福平杨俊李应国王国民保雨艾军张强李贤良
Owner INSPECTION & QUARANTINE TECH CENT OF CHONGQING ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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