CN-122012610-A - Application of corn biological clock gene Zmtoc b in improving drought tolerance of plants

Abstract

The invention is suitable for the fields of molecular biology and biotechnology, and provides application of a corn biological clock gene Zmtoc b in improving plant drought tolerance, wherein the nucleotide sequence of the corn biological clock gene Zmtoc b is shown as SEQ ID NO. 1, and the amino acid sequence of a protein Zmtoc1b encoded by the corn biological clock gene Zmtoc b is shown as SEQ ID NO. 2. The invention clones Zmtoc B gene from corn inbred line B73, transforms Arabidopsis through agrobacterium mediating method to obtain stable inheritance T3 generation transgene strain, drought stress treatment result shows that the drought tolerance of transgene Arabidopsis in seedling stage is obviously higher than wild type, the relative water content of leaf and SOD activity are raised, thus proving corn biological clock gene Zmtoc B can effectively improve plant drought tolerance, and provides key gene and technical support for plant stress tolerance improvement.

Inventors

• LI SHIPENG
• LEI ZIJIAN
• REN ZIHAN
• SU SHENGZHONG

Assignees

• 吉林大学

Dates

Publication Date

20260512

Application Date

20260403

Claims (4)

1. 1. The application of the corn biological clock gene Zmtoc b in improving plant drought tolerance is characterized in that the nucleotide sequence of the corn biological clock gene Zmtoc b is shown as SEQ ID NO. 1, and the amino acid sequence of the protein Zmtoc b encoded by the corn biological clock gene Zmtoc b is shown as SEQ ID NO. 2.
2. 2. Use of the maize biological clock gene Zmtoc b according to claim 1 to improve drought tolerance in plants, comprising the steps of: Soaking the non-flowering buds of the arabidopsis thaliana in agrobacterium tumefaciens bacteria solution containing Zmtoc b genes by a flower dipping method, normally culturing the soaked arabidopsis thaliana to maturity, harvesting T0 generation seeds, screening the T0 generation seeds by Basta, identifying by PCR, obtaining T1 generation transgenic Zmtoc b gene plants, and obtaining T3 generation transgenic Zmtoc b gene arabidopsis thaliana plants by twice generation adding.
3. 3. The use of the maize biological clock gene Zmtoc b according to claim 2 for improving drought tolerance in plants, wherein the method of constructing agrobacterium containing the Zmtoc1b gene comprises the steps of: Constructing an entry vector, namely connecting a target gene Zmtoc b to a vector pDONR207 to obtain a pDONR207-Zmtoc b plasmid, and connecting the pDONR207-Zmtoc b plasmid to an expression vector pEarley gate101 through an LR reaction to obtain a recombinant plasmid pEarley gate101-Zmtoc1b; Agrobacterium transformation recombinant plasmid pEarley Gate101-Zmtoc b was transformed into competent cells of Agrobacterium GV3101 to obtain Agrobacterium strain containing pEarley gate101-Zmtoc b.
4. 4. The use of corn biological clock gene Zmtoc b according to claim 3 for increasing plant drought tolerance, wherein in the step of portal vector construction, the LR response system is pDONR207-Zmtoc b plasmid 3 μl, pearley gate101 empty plasmid 1 μl, LR ClonaseTM II Enzyme mix μl, and total volume 5 μl.

Description

Application of corn biological clock gene Zmtoc b in improving drought tolerance of plants Technical Field The invention belongs to the field of molecular biology and biotechnology, and particularly relates to application of a corn biological clock gene Zmtoc b in improving plant drought tolerance. Background Corn (Zea mays l.) is an important food, feed and industrial feedstock crop. Corn belongs to gramineous crops with large water demand, the whole growth period of the corn is extremely sensitive to water stress, and drought is one of main abiotic stress factors limiting corn production, and the growth, development and yield formation of the corn are seriously affected. Therefore, the drought resistance of the corn is improved by molecular breeding by excavating and utilizing drought resistance gene resources, and the method has important significance for guaranteeing the grain safety. The method comprises the steps that the toc1 (Timing of CAB Expression 1) is an important component part in a biological clock core oscillator, belongs to the pseudoo-Response Regulator (PRR) family, and is named as the fact that the toc1 is used for regulating and controlling the expression of a CAB (Chlorophyll a/b-binding protein) gene, the CAB gene codes a key protein of a light-capturing compound and directly influences photosynthetic efficiency, and the circadian rhythm expression of a photosynthesis related gene is accurately controlled by directly combining with a CAB gene promoter or indirectly regulating and controlling related transcription factors, so that the energy capturing efficiency of plants in an illumination period is optimized. In arabidopsis thaliana, toc1 forms a negative feedback regulation loop with factors such as CCA1/LHY, and the like, so that the operation of a biological clock is accurately regulated, the biological processes such as photoperiod regulation, flowering time, hypocotyl elongation and the like are influenced, the realization of the function of the toc1 is closely related to the phosphorylation modification of the biological clock, and the latest research shows that the phosphorylation of the toc1 can obviously enhance the binding capacity of the biological clock with chromatin, so that the rhythmic expression of a downstream gene is regulated within a specific time window. In recent years, more and more researches show that the biological clock has close relation with plant adversity adaptation, the biological clock can regulate and control the rhythmic expression of antioxidant enzyme genes, influence the active oxygen scavenging capability of plants at different time points, meanwhile, the biological clock genes also participate in the regulation of ABA signal paths, influence the air pore opening and closing and the water utilization efficiency, and particularly, the latest researches show that core biological clock regulating factors (including toc1, PRRs and the like) can directly inhibit the expression of autophagy related genes (ATGs) in daytime, and activate the genes at night so as to match autophagy flows with the energy metabolism rhythms of cells, and the biological clock gating mechanism is an important strategy for adapting plants to environmental stress, thus providing a brand-new theoretical view for understanding the regulation drought resistance of the biological clock genes. The homologous gene of the toc1 in the corn is named Zmtoc b, and researches show that the gene presents rhythmic expression under different photoperiod conditions, participates in the photoperiod sensitivity regulation of the corn and is obviously related to the male-pulling period (BTA) and the silking period (BTL), however, the function of the toc1b gene in the corn in drought resistance has not been reported in detail. Disclosure of Invention The embodiment of the invention aims to provide an application of a corn biological clock gene Zmtoc b in improving plant drought tolerance, aiming at solving the problems in the background technology. In order to achieve the above purpose, the present invention provides the following technical solutions: The nucleotide sequence of the corn biological clock gene Zmtoc b is shown as SEQ ID NO. 1, and the amino acid sequence of the protein Zmtoc1b encoded by the corn biological clock gene Zmtoc1b is shown as SEQ ID NO. 2. Compared with the prior art, the invention has the following specific beneficial effects: The invention clones Zmtoc B gene from corn inbred line B73, transforms Arabidopsis through agrobacterium mediating method to obtain stable inheritance T3 generation transgene strain, subcellular localization result shows Zmtoc B protein is localized in cell nucleus, drought stress treatment result shows that drought tolerance of transgene Arabidopsis in seedling stage is obviously higher than wild type, leaf relative water content and SOD activity are raised, thereby proving corn biological clock gene Zmtoc B can effectively improve drought tol