Foot-and-mouth disease virus Asia I infectious cDNA and construction method thereof

A technology of foot-and-mouth disease virus and construction method, which is applied in the direction of recombinant DNA technology, botany equipment and method, biochemical equipment and method, etc., and can solve problems such as restricting pathogenic mechanism

Inactive Publication Date: 2008-07-23
云南省畜牧兽医科学研究所
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  • Claims
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AI Technical Summary

Problems solved by technology

At present, the infectious cDNA molecule of Asian type 1 foot-and-mouth disease virus has not been successfully constructed at home and abroad, which seriously restricts the development of many fields such as the pathogenic mechanism, molecular immune mechanism and new vaccine research of Asian type 1 foot-and-mouth disease virus.

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  • Foot-and-mouth disease virus Asia I infectious cDNA and construction method thereof

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Embodiment

[0064] The 5' end upstream of the Asia-1 foot-and-mouth disease virus infectious cDNA provided by the present invention has a T7 RNA polymerase promoter, and the 5' non-coding region of the genome has a Poly (C) sequence, and the Poly (C) sequence has 2 to 50 C, It is preferably 10-40 C, the best is 12-36 C, and the 3' end has a single restriction endonuclease site and a Poly(A) sequence at the tail, and the Poly(A) sequence has 18-70 A, preferably 18-50 A, most preferably 30-36 A, the difference between the infectious cDNA and the natural sequence of the viral genome is no more than 1-30 nucleotides, preferably no more than 1-20 nucleotides , preferably no more than 1 to 10 nucleotides.

[0065] The infectious cDNA constructed by the present invention is to clone the full-length cDNA of the Asian type 1 foot-and-mouth disease virus genome into a plasmid vector, which includes Poly(C) and Poly(A) required to ensure that the synthesized transcript has infectivity sequence. Th...

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Abstract

The invention relates to an infectious cDNA of foot-and-mouth disease virus of type Asia 1 and the construction method, wherein, the RNA transcribed from the infectious cDNA contains the host cell of the infectious cDNA. The construction method is as follow: the foot-and-mouth disease virus of type Asia 1 is used as template; the genome segments are expanded by reverse transcription using specific primer; the expanded segments are assembled and cloned to a carrier; T7RNA polymerized polymerase promoeter is positioned at the upper reach of the obtained 5' end of cDNA; the 3' end is provided with single restriction enzyme cutting site and tail end Poly (A) sequence; the difference between the infectious cDNA and virus genome natural sequence is less than thirty nucleotides. The infectious cDNA of foot-and-mouth disease virus of type Asia 1 can be used to research the pathogenesis of virus and the replication and the expression and regulation mechanism of genome, and also can be used to prepare marked attenuated strain vaccine and develop new foot-and-mouth disease vaccine with gene manipulation technique.

Description

technical field [0001] The invention relates to a foot-and-mouth disease virus infectious cDNA and a preparation method thereof, in particular to a method for constructing an Asian type-1 foot-and-mouth disease virus ZB / CHA / 58(att) infectious cDNA. Belongs to the field of biotechnology. Background technique [0002] The establishment of infectious cDNA technology has become one of the most important tools in modern virology research. Researchers can effectively use the full-length cDNA of the virus as a template to transcribe infectious RNA in vitro to recover RNA viruses. The core of this technology is to firstly construct the full-length cDNA molecule of RNA virus, make it under the control of RNA polymerase promoter, obtain viral RNA again through in vitro transcription process, and then transfect the transcript RNA into mammalian cells to rescue the live cells. Viruses, since this rescued virus is derived from a full-length cDNA molecule, various modifications or modifi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/42C12N15/09
Inventor 信爱国张念祖李华春杨永钦李乐
Owner 云南省畜牧兽医科学研究所
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