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Tobacco AKT2/3 gene and application

A gene and tobacco technology, applied in the field of tobacco AKT2/3 gene and its application, can solve the problems of unknown Shaker function and large potassium consumption in tobacco

Active Publication Date: 2019-02-19
GUIZHOU TOBACCO SCI RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Tobacco is a crop that consumes a lot of potassium. The potassium content of tobacco leaves is an important indicator to measure the quality of tobacco leaves. At present, there are few studies on potassium ion channels in tobacco, and the function of tobacco Shaker is unknown.

Method used

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  • Tobacco AKT2/3 gene and application
  • Tobacco AKT2/3 gene and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Cloning of Example 1 Tobacco AKT2 / 3 Gene

[0032] Get 0.5g of tobacco (tobacco variety is K326) fresh leaves, use the Trizol method to extract the total RNA of tobacco cells, then use the cDNA synthesis kit of TaKaRa company to synthesize cDNA, and further use Primer5.0 software to design and obtain primers through artificial optimization. The primers include a forward primer and a reverse primer, and the nucleotide sequence of the forward primer is: 5'-ATGCAAATGAGCCATTCATC-3'; the nucleotide sequence of the reverse primer is 5'-CTACAAGCAATTTGGATCT-3', Use the synthesized cDNA as a template for PCR amplification. The PCR amplification system is 20 μL, including 10 μL of Premix ExTaq, 0.5 μL of 10 μM forward primer, 0.5 μL of 10 μM reverse primer, 1 μL of tobacco cell cDNA, ddH 2 O 8 μL; the reaction program of the PCR amplification is: pre-denaturation at 95° C. for 5 minutes; denaturation at 95° C. for 30 seconds; annealing at 55° C. for 30 seconds; extension at 72° C....

Embodiment 2

[0034] Example 2 Biological Functional Analysis of Tobacco AKT2 / 3 Gene

[0035] 1. Purpose of the experiment

[0036] The biological function of tobacco AKT2 / 3 gene was verified by yeast functional complementation experiment.

[0037] 2. Experimental method

[0038] The potassium uptake-deficient yeast mutant strain R5421 was used as the recipient strain. Strain R5421 can be found in Maathuis F J Mand Sanders D 1996 Mechanisms of potassium absorption by higher plantroots. Physiol. Plant. 96, 158–168.

[0039] The T-vector connected with the tobacco AKT2 / 3 gene of Example 1 and the expression vector P416 (yeast free type shuttle expression vector, TEF constitutive promoter, CYC1 terminator, CEN6 ARSH4 origin of replication, the selection marker in yeast is URA3, large intestine The screening marker in Bacillus is Amp. Carrier P416 can be found in Functional Expression of aω-3Fatty Acid Desaturase Gene from Glycine max in Saccharomyces cerevisiae) to carry out double enzyme d...

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Abstract

The invention relates to a tobacco AKT2 / 3 gene and an application. The tobacco AKT2 / 3 gene and the coded protein sequence are shown as SEQ ID NO: 1 and 2 respectively. The AKT2 / 3 gene is cloned from tobacco for the first time, and the biological function of the AKT2 / 3 gene is verified through a complementary experiment of yeast function. The AKT2 / 3 gene can promote the absorption and transport ofpotassium ions.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to the tobacco AKT2 / 3 gene and its application. Background technique [0002] Potassium ion channels are ion channels that allow potassium ions to specifically permeate the plasma membrane, while hindering the permeation of other ions, especially sodium ions. These channels are generally composed of two parts: one part is the channel region, which selects and allows potassium ions to pass through, while blocking sodium ions; the other part is a gating switch, which switches the channel on and off according to the signal in the environment. [0003] In the prior art, studies on potassium ion channel genes are extensive in the model plant Arabidopsis thaliana. For example, the plant Shaker ion channel is structurally composed of four subunits arranged around a central domain. The hydrophobic region of each subunit is composed of 6 transmembrane segments (Trans-membrane se...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/00A01H6/82
CPCC07K14/415C12N15/8243
Inventor 余婧任学良李立芹鲁黎明张洁王仁刚郭玉双
Owner GUIZHOU TOBACCO SCI RES INST
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