Application of grape circSIZ1 in regulation and control of plant growth and development and salt stress resistance

A plant growth and grape technology, applied in the field of grape circSIZ1 in regulating plant growth and development and salt stress resistance, to achieve the effect of great application value

Active Publication Date: 2020-09-29
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] circRNA is a new hotspot in the field of non-coding RNA research, but there is no report on grapevine circRNA in regulating plant growth and development and salt stress resistance

Method used

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  • Application of grape circSIZ1 in regulation and control of plant growth and development and salt stress resistance
  • Application of grape circSIZ1 in regulation and control of plant growth and development and salt stress resistance
  • Application of grape circSIZ1 in regulation and control of plant growth and development and salt stress resistance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1: Cloning of grape circSIZ1

[0052] 1. Acquisition of plant material:

[0053] The plant material used in this experiment is the tissue culture seedling of Eurasian grape 'Cresson Seedless'. The experimental materials were cultivated in the artificial climate chamber of the College of Horticulture, Shandong Agricultural University. The culture medium is MS medium containing 30g / L sucrose and 6g / L agar. Grape leaves were collected for RNA extraction.

[0054] 2. RNA extraction and reverse transcription:

[0055] Use the MiniBEST Universal RNA Extraction Kit (TaKaRa) kit to extract grape total RNA according to the standard instructions. The integrity was detected by ordinary agarose gel electrophoresis (gel concentration 1.5%; 0.5×TAE electrophoresis buffer; 100v, 20min). The maximum rRNA brightness in the electrophoresis strip should be 1.5-2.0 times the brightness of the second rRNA, otherwise it indicates the degradation of the rRNA sample. RNA with goo...

Embodiment 2

[0062] Example 2: Expression of grape circSIZ1 in different tissues and different salt treatment time points

[0063] 1. Acquisition of plant material:

[0064] On June 2, 2020, grape roots, stems, leaves, flowers, and fruits were collected at the grape experiment base of Shandong Agricultural University. The samples were wrapped in aluminum platinum paper and immediately put into liquid nitrogen. After returning to the laboratory, RNA was extracted.

[0065] 'Creasen Seedless' tissue culture seedlings, the medium is MS basic medium plus indole butyric acid (final mass concentration is 0.2mg / L), the culture temperature is (25±1)°C, and the light intensity is 2000-2400lx , Light and dark cycle 14h / 10h. The grape tissue-cultured seedlings subcultured for one month and with basically the same growth were randomly divided into treatment group and control group, with 9 plants in each group; 3 replicates were set up in the treatment group and control group, with 3 plants in each re...

Embodiment 3

[0083] Example 3: Functional verification of grape circSIZ1 transfected with tobacco

[0084] The transformation vector was pHB, and the circSIZ1 overexpression vector was constructed ( Figure 4). The construction process is to clone the circSIZ1 loop-forming region from DNA, plus 500 bp upstream and downstream of its flanking intron, and then connect it to the middle of a specific intron and its reverse complementary intron; the specific construction method of the circSIZ1 overexpression vector can be found in the patent "A plant circRNA overexpression vector and its construction method" (application number: CN201910199215.3). The leaf disk method was used to transform wild-type tobacco to obtain transgenic lines with overexpression of circSIZ1.

[0085] First, pipette 5 μL of Agrobacterium containing the circSIZ1 overexpression vector and inoculate it into a Erlenmeyer flask containing kanamycin and rifampicin LB culture solution, and activate it at 28°C and 200 rpm for 2...

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Abstract

The invention discloses application of grape circSIZ1 to regulation and control of plant growth and development and salt stress resistance, and belongs to the technical field of biology. The circSIZ1provided by the invention is circRNA derived from a SUMO E3 ligase gene, the full length of the circSIZ1 is 433bp, and the nucleotide sequence of the circSIZ1 is shown as SEQ ID NO. 1. And the expression quantity of circSIZ1 after salt stress treatment is in a rising trend. The circSIZ1 is overexpressed in tobacco, so that the growth and development of a root system are promoted, and in addition,the salt resistance of a transgenic plant is remarkably improved. The invention provides a theoretical basis for improving the stress resistance of plants by utilizing a gene engineering technology inthe future, and has a great application value.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the use of grape circSIZ1 in regulating plant growth and development and salt stress resistance. Background technique [0002] Circular RNA (circular RNA, circRNA) refers to a covalently closed circular RNA molecule formed by trans-splicing. The combination of high-throughput sequencing technology and bioinformatics tools has revealed that circRNAs are ubiquitous in a variety of organisms. circRNA has the characteristics of high stability, sequence conservation and tissue specificity. In the past five years, reports on the identification of circRNAs in plants have involved more than 20 plant species, including monocots rice, maize, barley, wheat, sorghum, millet, Brachypodium distachyon, Moso bamboo and Ganoderma lucidum, dicotyledons Plants Arabidopsis, tobacco, pepper, tomato, cabbage, potato, soybean, peanut, kiwi, citrus, pear, grape, orange, sea buckthorn, poplar, cotton and t...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12N15/82A01H5/00A01H6/82
CPCC12N15/113C12N15/8273C12N15/8261C12N15/8216C12N2310/532
Inventor 高振孙宝箴杜远鹏姚玉新翟衡郑成超
Owner SHANDONG AGRICULTURAL UNIVERSITY
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