Preparation, detection and application of polyclonal antibody of Yam mild mosaic virus

A technology of polyclonal antibody and mosaic virus, applied in the field of genetic engineering, can solve the problems of production loss of yam, affecting the yield and quality of yam

Active Publication Date: 2014-01-29
GUANGXI UNIV
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  • Abstract
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Problems solved by technology

Viral diseases seriously affect the yield and quality of yam, causing heavy losses in the production of yam

Method used

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  • Preparation, detection and application of polyclonal antibody of Yam mild mosaic virus
  • Preparation, detection and application of polyclonal antibody of Yam mild mosaic virus
  • Preparation, detection and application of polyclonal antibody of Yam mild mosaic virus

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Embodiment Construction

[0046] Materials used include: HS high-fidelity DNA polymerase and T4 DNA ligase were purchased from TaKaRa Company. Column type DNA gel extraction kit (E.Z.N. Gel Extraction Kit) was purchased from OMEGA Company. RNAplant plus Total Plant RNA Extraction Reagent and Plasmid DNA Extraction Kit were purchased from Tiangen Biochemical Technology (Beijing) Co., Ltd., 5’ / 3’RACE Kit and PCR Product Purification Kit were purchased from Roche Company. TMB substrate chromogenic kit and protein extraction kit were purchased from Kangwei Century Company. Agarose is Regular Agarose G-10. Tryptone, yeast extract, urea, BSA, PMSF, TEMED, and DTT were purchased from Sigma. Restriction enzymes, RNase Inhibitor, IPTG, RevertAid TM First-strand cDNA synthesis kit, dNTP, GeneRuler1kb DNA ladder, GeneRuler1kb plus DNA ladder, PageRuler Prestained Protein Ladder, PageRuler Unstained Protein Ladder were purchased from Thermo Scientific. Ampicillin (Amp), Kanamycin (Kan), RNaseA, Acrylamid...

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Abstract

The invention discloses preparation, detection and application of a polyclonal antibody of a Yam mild mosaic virus (YMMV). Based on a YMMV genome, the inventor purifies a nuclear inclusion protein/coat protein (NIb/CP) of the YMMV through prokaryotic expression and uses the nuclear inclusion protein/coat protein to prepare the polyclonal antibody for an immune rabbit. The polyclonal antibody can specifically identify the Nib/CP protein of YMMV through prokaryotic expression and the Nib/CP protein of the YMMV virus which infects common yam rhizome, and generates specific immunity response. On the basis, the inventor further builds a set of efficient, high sensitive and accurate IC-RT-PCR (Immunocapture-Reverse Transcription-Polymerase Chain Reaction, ELISA (Enzyme-Linked Immuno Sorbent Assay) and Western blot methods which can specifically detect the YMMV from a common yam rhizome plant which is infected by YMMV. By applying the invention, a material basis and a technical support can be provided for research on interaction of the YMMV and the host, quick detection of the virus and somatotype and molecular biology study, thereby laying a solid foundation for epidemiologically monitoring and preventing treating the virus.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to prokaryotic expression of NIb / CP protein of yam mild mosaic virus, preparation, detection and application of polyclonal antibody. Background technique [0002] Yam, also known as Chinese yam and Chinese yam, is a plant of the family Dioscorea. Its roots are rich in starch, protein, vitamins, amino acids and various medicinal ingredients such as choline, saponins, mucopolysaccharides, etc., and also contain Fe, Zn, Cu, Ca, etc. A variety of minerals, depending on the species and varieties, can be used for medicine or food, with good market prospects and industrial development potential. Viral diseases have seriously affected the yield and quality of Chinese yam, and caused great losses in the production of Chinese yam. The literature shows that the viruses that infect yam under natural conditions include Dioscorea alata bacilliform virus (DaBV), Yam mosaic...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/08C07K16/06G01N33/569G01N33/543G01N33/68C12Q1/70C12Q1/68C12R1/94
Inventor 陈保善邹承武蒙姣荣张蕾卢晓静
Owner GUANGXI UNIV
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