Novel cinnamomum hupehanum ribosome inactivated protein gene CcRIPII

A ribosome inactivation and genetic technology, applied in genetic engineering, plant genetic improvement, microbial measurement/inspection, etc., can solve the problems of no successful reports on woody plant genetic transformation, etc., to achieve appropriate size, convenient detection, and high efficiency high effect

Inactive Publication Date: 2013-07-03
泰安市泰山林业科学研究院,翁曼丽 +1
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Problems solved by technology

However, the genetic transformation of woody plants has not been successfully reported so far

Method used

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  • Novel cinnamomum hupehanum ribosome inactivated protein gene CcRIPII
  • Novel cinnamomum hupehanum ribosome inactivated protein gene CcRIPII
  • Novel cinnamomum hupehanum ribosome inactivated protein gene CcRIPII

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Experimental program
Comparison scheme
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Embodiment Construction

[0010] (1) The in vitro regeneration system of woody plants is used as the transformation material, and the genetic transformation is carried out by the dipping method of Agrobacterium.

[0011] (2) Conversion operation steps

[0012] 1. Agrobacterium transformation method

[0013] (1) The pCAMBIA2300-CcRIPII expression vector was transformed into Agrobacterium tumefaciens LBA4404 by triparental strain hybridization method for the Agrobacterium dipping transformation experiment.

[0014] (2) Bacterial liquid preparation

[0015] 1) Use an inoculation loop to scrape the surface of the frozen culture of Agrobacterium strains containing recombinant plasmids, streak on the above-mentioned YEP plate containing the corresponding antibiotics, and place it in a constant temperature incubator at 28°C for 2 days, until colonies grow out on the plate It can be used for liquid culture. Plates with colonies were stored in a 4°C refrigerator. After one month, transfer to a new YEP plate...

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Abstract

The invention designs a PCR (Polymerase Chain Reaction) primer according to a conserved sequence of a reported plant ribosome inactivated protein gene CcRIPII, and obtains a novel partial sequence of a novel cinnamomum hupehanum ribosome inactivated protein gene CcRIPII through homologous amplification; subsequently a full-length gene cDNA (Complementary Deoxyribonucleic Acid) sequence is obtained through RACE (Rapid-Amplification of Complementary Deoxyribonucleic Acid Ends); the CcRIPII gene does not contain internal functors but contains a complete ORF (Open Reading Frame) area; and the ORF contains full-length 1758 nucleotides, coding 585 amino acids and one termination codon (TGA). A plant expression carrier pCAMBUA2300-CcRIPII of the CcRIPII gene is established, and a transgenetic seedling is obtained by using pCAMBIA2300-CcRIPII transformed Ningyang Chinese-date. The CcRIPII is the first woody plant ribosome inactivated protein gene which is cloned and subjected to wood genetic transformation on the international.

Description

1. Technical field [0001] The invention relates to a new ribosome inactivation protein gene CcRIPII of camphor tree. The CcRIPII gene has no intron, but contains a complete ORF region. Its coding sequence is 1758 nucleotides (29-1786) in length, encoding 585 amino acids and a stop codon (TGA). [0002] The plant expression vector pCAMBIA2300-CcRIPII of CcRIPII gene was constructed, and the transgenic plants were obtained by transforming Ningyang jujube with pCAMBIA2300-CcRIPII. CcRIPII is the first ribosome-inactivating protein gene of woody plants that has been cloned and genetically transformed into forest trees in the world. Its cloning and success in genetic transformation of forest trees are of great significance to the research of anti-virus and genetic breeding of woody plants. 2. Background technology [0003] Ningyang jujube in Shandong is a famous local specialty in my country. In recent years, the prevalence of viral diseases has seriously affected the developmen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/84A01H5/00C12Q1/68C12N15/11
Inventor 冯殿齐翁曼丽刘静王斌赵进红罗磊黄艳艳张虹
Owner 泰安市泰山林业科学研究院,翁曼丽
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