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Tobacco OA1 gene, and primer and application thereof

A tobacco and gene technology, applied in the application, genetic engineering, plant genetic improvement and other directions, can solve the problems of insufficient oil content of tobacco raw materials, restricting the application of high-quality tobacco leaves, etc., and achieve the effects of huge economic potential, high expression, and broad application prospects.

Active Publication Date: 2020-02-04
河南省烟草公司三门峡市公司 +1
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Although China's tobacco leaf production level has been greatly improved, the problem of insufficient oil content in tobacco leaf raw materials is very prominent, which largely restricts the application of high-quality tobacco leaves in high-end cigarette formulations

Method used

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  • Tobacco OA1 gene, and primer and application thereof
  • Tobacco OA1 gene, and primer and application thereof
  • Tobacco OA1 gene, and primer and application thereof

Examples

Experimental program
Comparison scheme
Effect test

experiment example 1

[0024] Experimental Example 1 Comparison of Tobacco Leaf Oil Content in Different Tobacco Varieties (Lines)

[0025] The tested varieties (lines) were Yuyan 6, Qinyan 96, Yunyan 87, China Tobacco 100, 89112, X101, X126, X131 (planted in Xuchang Science and Education Park in 2017-2018). The soil for testing is cinnamon soil, the organic matter content in the soil is 9.96g / kg, the total nitrogen content is 0.95g / kg, the alkaline nitrogen content is 73.54mg / kg, the soil available phosphorus content is 34.12mg / kg, and the available potassium content is 130.70 mg / kg.

[0026] When the first central flower of the tobacco plant is 50% open, topping is carried out, and the number of leaves per plant is 22, and the field management is carried out according to the production technology of high-quality flue-cured tobacco. After the tobacco leaves were rounded, 10 plants were established respectively. The tobacco leaves of the tobacco plants were from bottom to top, and the 17th to 22nd ...

experiment example 2O

[0030] Experimental example 2 Discovery and cloning of OA1 (OilAccumulation 1) gene

[0031] Transcriptome sequencing analysis was performed on the mature leaves of the high-oil content line X101 and the low-oil content variety Yunyan 87. Through the identification of differentially expressed genes, it was found that the abundance of a gene transcript in the high-oil line X101 was 6 times that of the low-oil line Yunyan 87. Compared with the existing database, no similar sequence and functional annotation information were found, and the gene was preliminarily determined to be a new gene, and the gene was named Oil Accumulation 1, OA1.

[0032] (1) First-strand cDNA synthesis of tobacco leaves

[0033] The isolation and purification of total RNA from mature leaves of tobacco high-oil content line X101 was carried out according to the instructions of the TRIZOL kit (ThermoFisher Scientific, USA). Absorb 1-2 μg of total RNA from tobacco leaves into a 1.5mL centrifuge tube, foll...

experiment example 3O

[0040] EXPERIMENTAL EXAMPLE 3 Expression analysis of OA1 gene in tobacco varieties (lines) with different oil content

[0041] Fluorescent quantitative PCR (QPCR) method was used to analyze the expression of OA1 gene in tobacco varieties (lines) with different oil content. The experimental samples were mature leaves of 8 tobacco varieties (lines) in Experimental Example 1. In each sample, according to the method in Experimental Example 2, 3 μg of total RNA was reverse-transcribed into first-strand cDNA as a template.

[0042] Design fluorescent quantitative PCR specific primers according to the full-length cDNA sequence of OA1 gene, as follows:

[0043] Upstream primer: 5'-GGAGTGACCGTGGTAGTTTG-3' (SEQ ID NO.5);

[0044] Downstream primer: 5'-CCAGTTAGGAGATGTGGCA-3' (SEQ ID NO. 6).

[0045] The amplification reaction was performed on an ABI real-time quantitative PCR instrument, and the amplification product was 144bp.

[0046] Each sample amplifies the tobacco Actin2 gene f...

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Abstract

The invention discloses a tobacco OA1 gene as well as a primer and application thereof. The tobacco OA1 gene has a nucleotide sequence shown as SEQ ID NO. 1. The tobacco OA1 gene disclosed by the invention is capable of affecting accumulation of oil in tobacco leaves and regulating oil content of the tobacco leaves, as well as high in expression in high-oil varieties; and thus, the tobacco OA1 gene has huge economic benefit potentials.

Description

technical field [0001] The invention relates to a tobacco gene, in particular to a tobacco OA1 gene, primers and applications. Background technique [0002] Oil is a soft liquid substance contained in tobacco leaf tissue cells, mainly including: volatile oil, resin, grease, fatty acid, wax, lipid, sterol, pigment, etc. Tobacco leaf oil content is an important basis for measuring the appearance quality of tobacco leaves. Studies have shown that the oil content of tobacco leaves has a significant positive correlation with sugar content, but a significant negative correlation with total nitrogen content. Insufficient oil content will lead to an uncoordinated sugar-alkaline ratio in tobacco leaves, thinner tobacco leaves, and poor hygroscopicity and combustibility. Tobacco leaves with a lot of oil content have a good aroma, sufficient aroma, lighter miscellaneous gas, and a purer and more comfortable aftertaste. Moreover, the higher the tobacco leaf oil content, the better its...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/82C12N15/11C12N5/10C07K14/415C12Q1/6851A01H5/12A01H6/82
CPCC07K14/415C12N15/8247C12Q1/6851C12Q2531/113C12Q2563/107
Inventor 苏新宏韦凤杰陈彦春夏宗良张小全李洪臣杨军杰杨晋燕
Owner 河南省烟草公司三门峡市公司
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