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A potassium transporter kup9 from tobacco and its coding gene and application

A gene and tobacco technology, applied in the field of potassium transporter KUP9 and its coding gene and application, can solve the problems of lack of absorption capacity, decreased survival rate, decreased potassium ion concentration, 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

[0003] Existing technology studies on potassium transporter genes are extensive in the model plant Arabidopsis thaliana. For example, studies have shown that Arabidopsis mutant kup6 has a lower survival rate under drought stress than wild-type plants, while 35S::KUP6 overexpression The drought tolerance of transgenic plants was significantly enhanced (Shabala et al., 2007); while the mutant seedlings of KUP7 gene exhibited a low-potassium-sensitive phenotype of yellowing leaves under low-potassium stress, and the potassium ions in the roots The content decreased significantly, and the loss of KUP7 gene function reduced the ability of Arabidopsis seedlings to absorb potassium under low potassium conditions, and the concentration of potassium ions in xylem juice decreased (Min et al., 2016)
[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 transporter genes in tobacco, and the function of tobacco KUP is unknown.

Method used

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  • A potassium transporter kup9 from tobacco and its coding gene and application
  • A potassium transporter kup9 from tobacco and its coding gene and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] The acquisition of embodiment 1 KUP9 gene

[0047] 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'-ATGACTTCAGGAATGGAAAT-3'; the nucleotide sequence of the reverse primer is 5'-TTACACATAAAAGATCTGTC-3', with the cDNA synthesized as a template , for PCR amplification.

[0048] 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.

[0049] 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.

[0050...

Embodiment 2

[0051] Example 2 The role of KUP9 gene in promoting the absorption and transport of potassium ions

[0052] The T-vector and the expression vector P416 connected with the KUP9 gene described in Example 1 were subjected to double enzyme digestion (restriction sites: Sma I and Xho I), and the target gene and expression vector P416 were recovered, and then ligated with Enzyme ligation, transfer the ligated recombinant yeast expression vector into Escherichia coli DH5α competent cells, perform PCR amplification and enzyme digestion on the transformed Escherichia coli colony to verify whether the construction is successful, and construct the successful recombinant yeast expression vector Transfer to R5421.

[0053] 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 the liquid ...

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Abstract

The invention relates to the field of genetic engineering, and specifically discloses a potassium transporter KUP9 from tobacco, its coding gene and application. The present invention provides for the first time the KUP9 gene isolated from tobacco. The full length of the KUP9 gene is 2370 bp. After functional verification, the KUP9 gene provided by the present invention is transferred into the potassium absorption-deficient yeast mutant strain R5421, and then expresses the recombinant yeast of the KUP9 gene. It regained the function of potassium ion absorption and transport. Therefore, the KUP9 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 KUP9 from tobacco and its coding gene and application. Background technique [0002] Potassium transporter is a carrier protein that can transport potassium ions into cells when the external potassium ion concentration is extremely low. Potassium transporters are usually some transmembrane proteins, which complete the transmembrane transport of potassium ions through conformational changes. It is an active transport process that requires ATP to provide energy, usually K + / H + or K + / Na + Collaborative transport. [0003] Existing technology studies on potassium transporter genes are extensive in the model plant Arabidopsis thaliana. For example, studies have shown that Arabidopsis mutant kup6 has a lower survival rate under drought stress than wild-type plants, while 35S::KUP6 overexpression The drought tolerance of transgenic plants was significa...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/81C12N1/19C12N15/82A01H5/00A01H6/82A01H6/20
CPCC07K14/415C12N15/81C12N15/8243
Inventor 曾吉凡任学良鲁黎明李立芹王仁刚张洁王自力郭玉双
Owner GUIZHOU TOBACCO SCI RES INST
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