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Tobacco kup1 gene and its application

A gene and tobacco technology, applied in the tobacco KUP1 gene and application fields, can solve the problems of large potassium consumption, reduced potassium ion concentration, and decreased

Active Publication Date: 2021-08-31
GUIZHOU TOBACCO SCI RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Existing technology studies on potassium transporter genes are extensive in the model plant Arabidopsis. 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 However, the drought tolerance of transgenic plants was significantly enhanced (Shabala et al., 2007); while the KUP7 mutant seedlings showed a low-potassium-sensitive phenotype of yellowing leaves under low-potassium stress, and the potassium ion content in the roots was significantly The loss of KUP7 gene function decreased the ability of Arabidopsis seedlings to absorb potassium under low potassium conditions, and decreased the concentration of potassium ions in xylem sap (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 gene is unknown.

Method used

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Examples

Experimental program
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Effect test

Embodiment 1

[0031] Cloning of Example 1 Tobacco KUP1 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, the nucleotide sequence of the forward primer is: 5'-ATGTATCAAGTAGATATTGA-3'; the nucleotide sequence of the reverse primer is 5'-CTAGACGTAGTATATCATGC-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. for ...

Embodiment 2

[0034] Example 2 Biological function analysis of tobacco KUP1 gene

[0035] 1. Purpose of the experiment

[0036] The biological function of tobacco KUP1 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 KUP1 gene in Example 1 and the expression vector P416 were subjected to double enzyme digestion (the restriction sites were XbaI and XhoI), respectively, and the target gene and expression vector P416 (yeast episomal shuttle expression vector, composed of TEF) were recovered. type promoter, CYC1 terminator, CEN6 ARSH4 origin of replication, the screening marker in yeast is URA3, and the screening marker in Escherichia coli is Amp. Vect...

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Abstract

The invention relates to tobacco KUP1 gene and its application. The sequences of the tobacco KUP1 gene and its encoded protein are shown in SEQ ID NO: 1 and 2, respectively. The present invention clones the KUP1 gene from tobacco for the first time, and verifies the biological function of the gene through the yeast function complementation experiment. The tobacco KUP1 gene has the function of promoting potassium ion absorption and transport.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to tobacco KUP1 gene and its 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 significantly enhanced (Shabala et al.,...

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 林世锋任学良李立芹鲁黎明耿召良王仁刚张洁
Owner GUIZHOU TOBACCO SCI RES INST
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