Mutant flue-cured tobacco CYP82E4 gene, and applications thereof

Abstract

The invention discloses a mutant flue-cured tobacco CYP82E4 gene. The nucleotide sequence is represented by SEQ ID NO.2; and when the mutant flue-cured tobacco CYP82E4 gene is compared with wild type CYP82E4 gene, nucleotide G at the 116th site is changed into A via mutation. The invention also discloses a protein encoded by the mutant flue-cured tobacco CYP82E4 gene, and the amino acid sequence of the protein is represented by SEQ ID NO.4. It is shown by research that the mutant flue-cured tobacco CYP82E4 gene is capable of reducing expression or functions of nicotine N-demethylase, inhibiting the activity of nicotine N-demethylase in mature leaves, and reducing nornicotine content of tobacco plants containing the mutant flue-cured tobacco CYP82E4 gene obviously. The mutant flue-cured tobacco CYP82E4 gene can be used in breeding and seed selection of tobacco varieties with reduced nornicotine content; and tobacco varieties with low nornicotine content can be obtained via selfing of tobacco plants containing the mutant flue-cured tobacco CYP82E4 gene, or conventional selection hybridization with other flue-cured tobacco.

Description

technical field [0001] The invention relates to a mutant flue-cured tobacco CYP82E4 gene, its encoded protein, and its application in reducing the expression or function of nicotine demethylase. Background technique [0002] Tobacco alkaloids mainly include nicotine, nornicotine, anatabine, and anatabine. These alkaloids generate NNK, NNN, NAT, and NAB respectively through direct or indirect nitrosation. Numerous animal experiments have shown that NNK and NNN are strong carcinogens, while NAB and NAT have only weak or no obvious carcinogenicity. When tobacco leaves are prepared and processed, the tobacco-specific nitrosamine (TSNA) compound N-nitrosonornicotine (NNN) produced by the nitrosation reaction of nornicotine may cause esophageal cancer and oral cancer (HechtSS Biochemistry, Biology, and Carcinogenicity of Tobacco- Specific N-Nitrosamines. Chemical Research in Toxicology 1998, 11: 559-603; Hecht SS. Tobaccocarcinogens, their biomarkers and tobacco-induced cancer. N...

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Problems solved by technology

[0005] Chinese patent CN102858983A has obtained the mutation of some sites of Burley tobacco CYP82E4 by non-transgenic method, resulting in the reduction of nornicotine and NNN content in Burley tobacco. Since more than 90% of the current tobacco leaf production in my country is flue-cured tobacco, the white The introduction of the mutation site of CYP82E4 in rib tobacco into flue-cured tobacco can be accomplished by crossing with flue-cured tobacco varieties, but crossing Burley tobacco with flue-cured tobacco will lead to poor quality of flue-cured tobacco. Therefore, in addition to creating new germplasm, in the process of breeding new flue-cured tobacco varieties, we must It is more difficult to transfer the beneficial traits of Burley tobacco to flue-cured tobacco without other chain burdens

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