Expression process of coat protein gene of prunus necrotic ring spot virus (PNRSV), antiserum and kit

Abstract

The invention discloses an expression process of a coat protein gene of prunus necrotic ring spot virus (PNRSV), an antiserum and a kit. The expression method comprises the following steps of: extracting a ribose nucleic acid (RNA) of the PNRSV in an affected material; adopting a transcription-polymerase chain reaction (RT-PCR) amplification process to clone the coat protein gene of the PNRSV; connecting the obtained coat protein gene to a pGEM-T vector, obtaining a pGEM-T-PNRSV recombinant vector, and using sequence analysis to determine the sequence accuracy of the coat protein gene of the PNRSV; building an eukaryotic expression vector of the coat protein gene of the PNRSV; transforming a pichia pastoris GS115 strain, and inducing a pichia pastoris expression coat protein; and using Western-blot to detect the size accuracy of an expression proteine. The invention aims to provide the process for using the pichia pastoris to express the PNRSV coat protein gene, the antiserum, and a PNRSV ELISA detection kit.

Description

technical field [0001] The invention relates to a method for expressing the coat protein gene of plum necrotic ringspot virus by using Pichia pastoris, and also relates to an antiserum prepared by using the expressed coat protein gene; the invention also relates to a plum necrotic ringspot virus ELISA Detection kit. Background technique [0002] Plum necrosis ringspot virus (PNRSV) is the latest quarantine pathogen of imported plants promulgated by my country. It is one of the most dangerous viruses for stone fruit trees. Once the fruit trees are infected by it, it will cause immature A large number of fruits fall off, the yield is severely reduced, and the quality of the fruit is greatly reduced. And the virus spreads fast, which may bring catastrophic losses to the development of the whole country's fruit tree industry in a short period of time. At present, there are only local reports of the virus in China. Since our country pursues the national policy of reform and ope...

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Problems solved by technology

However, due to the very low content of the virus in fruit trees, it is difficult to reproduce, so it becomes very difficult to purify the virus in large quantities and prepare high-titer antiserum, and the virus is currently in the list of quarantine viruses in my country, so it is not suitable for individual detection. The department provides the virus as a positive control, which largely limits serological testing for the virus

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