A potassium transporter tpk1 from tobacco and its coding gene and application

A gene, tobacco technology, applied to potassium transporter TPK1 and its encoding gene and application field, can solve the problems of unknown tobacco TPK function, large potassium consumption, etc., and achieve the effect of promoting potassium ion absorption and transport

Active Publication Date: 2021-08-31
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 TPK is unknown.

Method used

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  • A potassium transporter tpk1 from tobacco and its coding gene and application
  • A potassium transporter tpk1 from tobacco and its coding gene and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Obtaining of embodiment 1 TPK1 gene

[0046] Get 0.5g of fresh tobacco 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 the Primer5.0 software to design and obtain primers through artificial optimization. The primers include forward primers and a reverse primer, the nucleotide sequence of the forward primer is: 5'-ATGACTAACAGAGACAAAATCCAGC-3'; the nucleotide sequence of the reverse primer is 5'-TCACCTTCCTGTCCAAGACGATTGG-3', using the synthetic cDNA as a template , for PCR amplification.

[0047] The PCR amplification system is a 20 μL system, including: Premix ExTaq 10 μL, 10 μM forward primer 0.5 μL, 10 μM reverse primer 0.5 μL, tobacco cell cDNA 1 μL, ddH 2 08 μL.

[0048] The PCR amplification reaction program was: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 30 s, annealing at 55°C for 30 s, extension at 72°C for 2 min, and 35 cycles.

[0...

Embodiment 2

[0050] Example 2 The role of TPK1 gene in promoting the absorption and transport of potassium ions

[0051] The T-vector connected with the TPK1 gene described in Example 1 and the expression vector P416 were subjected to double enzyme digestion (restriction sites: SmaI and XhoI), and the target gene and expression vector P416 were recovered, and then ligated with ligase , transfer the ligated recombinant yeast expression vector into Escherichia coli DH5α competent cells, carry out PCR amplification and enzyme digestion on the transformed Escherichia coli single colony to verify whether the construction is successful, and transfer the successfully constructed recombinant yeast expression vector into into R5421.

[0052] The specific steps are as follows: Streak the preserved R5421 yeast on the solid medium YPDA with an inoculation loop, culture at 28°C for 12 hours; pick a single colony of R5421 yeast in the Ep tube, add 1mL of YPDA culture solution and vortex; Transfer all t...

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PUM

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Abstract

The invention relates to the field of genetic engineering, and specifically discloses a potassium transporter TPK1 from tobacco and its coding gene and application. The present invention provides for the first time the TPK1 gene isolated from tobacco. The full length of the TPK1 gene is 1062bp. After functional verification, the TPK1 gene provided by the present invention is transferred into the potassium absorption-deficient yeast mutant strain R5421, and the recombinant yeast expressing the TPK1 gene It regained the function of potassium ion absorption and transport. Therefore, the TPK1 gene provided by the present invention has the function of promoting the absorption and transport of potassium ions.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a potassium transporter TPK1 from tobacco and its coding gene and application. Background technique [0002] Potassium ion channels are ion channels that allow potassium ions to specifically permeate the plasma membrane, while blocking 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, studies have shown that the TPK / KCO channel in the model plant Arabidopsis includes two-pore potassium channel TPK (also known as tandem pore channel) and a Kir -type channel. T...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/00A01H6/20A01H6/82
CPCC07K14/415C12N15/8243
Inventor 任学良王仁刚鲁黎明李立芹郭玉双张洁王自力林世锋
Owner GUIZHOU TOBACCO SCI RES INST
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