Application of gene VLNHX3D in regulation of Na<+> and/or K<+> concentration of plant cells

A plant cell and transgenic plant technology, applied in botany equipment and methods, angiosperms/flowering plants, applications, etc., can solve problems such as the unknown state of gene function, achieve high salt tolerance, and improve plant salt tolerance.

Pending Publication Date: 2022-02-11
HENAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] Today, although the genome sequence of cotton has been sequenced, the functions of some genes are still unknown and require further exploration by researchers

Method used

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  • Application of gene VLNHX3D in regulation of Na&lt;+&gt; and/or K&lt;+&gt; concentration of plant cells
  • Application of gene VLNHX3D in regulation of Na&lt;+&gt; and/or K&lt;+&gt; concentration of plant cells
  • Application of gene VLNHX3D in regulation of Na&lt;+&gt; and/or K&lt;+&gt; concentration of plant cells

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] 1. Gene cloning

[0074] 1. Extraction of total RNA: Take the roots, stems and leaves of cotton materials frozen at -80°C and grind them into powders in liquid nitrogen, respectively, and put 100 mg of sample powders into 2 mL RNase-Free centrifuge tubes. Subsequent experimental steps were carried out according to the instructions of Tiangen RNA Extraction Kit (DP441).

[0075] 2. cDNA synthesis: cDNA was synthesized using the reverse transcription reagent HiScript III 1st Strand cDNA Synthesis Kit (+gDNA wiper) (R312-01) from Novozyme.

[0076] 3. Gene cloning (RT-PCR): using the first-strand cDNA obtained by reverse transcription as a template, the high-fidelity enzyme KOD-Plus-Neo (Code No. KOD-401) from TaKaRa Company was used to amplify the target gene.

[0077] Table 2 Gene Cloning PCR System

[0078]

[0079] Table 3 Gene Cloning PCR Reaction Program

[0080]

[0081] 5. At the end of the PCR reaction, use 1.5% agarose gel electrophoresis to detect the P...

Embodiment 2

[0111] 1. Real-time fluorescent quantitative PCR of VLNHX3D

[0112] 1. Cotton materials are roots, stems and leaves at various time points after salt treatment, and RNA is extracted.

[0113] 2. cDNA synthesis: HiScriptII Q RT SuperMix for qPCR (+gDNAwiper) (R233-01) reverse transcription reagent from Novizyme was used.

[0114]3. Design specific fluorescent quantitative primers: use primerpremier 5 to design specific primers for VLNHX3D and GhNHX3A, GhHIS3 is used as an internal reference gene, and the primer sequences are shown in Table 1.

[0115] 4. Real-time fluorescent quantitative PCR (RT-qPCR): use ChamQ Universal SYBR qPCR MasterMix to prepare the following reaction mixture.

[0116] Three biological replicates were performed for each sample, and the results were obtained using 2 -△Ct Perform analytical calculations.

[0117] Among them, the expression level of VLNHX3D in the leaves of upland cotton under no salt stress treatment (0, 1, 3, 6 and 12h) was used as t...

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PUM

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Abstract

The invention relates to the field of plant molecular biology, in particular to application of a gene VLNHX3D to regulation of Na<+> and / or K<+> concentration of plant cells. The invention provides the application of a gene VLNHX3D in regulating the Na<+> and / or K<+> concentration of plant cells. The gene VLNHX3D has a nucleotide sequence as shown in SEQ ID NO: 1. After the expression of the VLNHX3D is reduced by cloning the gene VLNHX3D, constructing a transgenic vector and utilizing a VIGS technology, under a salt stress condition, the Na<+> accumulation amount in roots, stems and leaves of a silent plant is remarkably increased, the K<+> content in the roots is remarkably reduced, and the salt tolerance of cotton is remarkably reduced, so that the VLNHX3D participates in early salt stress response, and the salt tolerance of the cotton is improved by regulating steady-state balance of Na<+> and K<+> in cells.

Description

technical field [0001] The present invention relates to the field of plant molecular biology, in particular, relates to gene VLNHX3D in regulating plant cell Na + and / or K + Concentration applications. Background technique [0002] Salt stress can lead to stunted growth and development of plants, yellowing or withering of leaves, and in severe cases, the whole plant will dry up and die. The harm of salt stress mainly includes three aspects: cell osmotic stress, ion toxicity and nutritional imbalance. [0003] Osmotic stress: When plants are subjected to salt damage, the original water balance of plant cells is broken. The water absorption capacity of plants is reduced, which leads to physiological drought of plants, resulting in inhibition of plant production. Osmotic stress rapidly reduces cell expansion in root tips and young leaves, resulting in stomatal closure. The increase of sodium ion content in the soil not only rapidly reduced soil water availability, but also...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/82C12Q1/6895C07K14/415A01H5/00A01H6/20A01H6/46A01H6/54A01H6/60A01H6/82
CPCC07K14/415C12N15/8273C12Q1/6895C12Q2600/13C12Q2600/158
Inventor 刘伟马宗斌马兴立朱伟
Owner HENAN AGRICULTURAL UNIVERSITY
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