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A longan flowering regulation gene dlerf23 and its protein and application

A technology of flowering regulation and protein regulation, which is applied in the fields of application, genetic engineering, plant gene improvement, etc., and can solve the problems of not having longan flowering, fruit farmers increasing production but not increasing income, etc.

Active Publication Date: 2021-07-23
重庆圣盈达科技开发有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In my country's main production areas, such as the western part of Guangdong and Hainan, because they do not have the low temperature required for longan flowering, it is often necessary to use KClO 3 To urge flowers, but this method has great limitations, such as being restricted by species, environment and other factors [3]
At the same time, the centralized listing of longan also often results in the phenomenon of increasing production but not increasing income of fruit farmers.

Method used

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  • A longan flowering regulation gene dlerf23 and its protein and application
  • A longan flowering regulation gene dlerf23 and its protein and application
  • A longan flowering regulation gene dlerf23 and its protein and application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The cloning of embodiment 1 target gene

[0027] Materials and methods

[0028] 1.1 Plant material

[0029] Three groups of 'Sijimi' longan and the main cultivar 'Shixia' longan with the same growth vigor and tree age (9 years old) were selected as sampling trees, and 'Shixia' longan showed seasonal flowering as other cultivars. , SF). Its flowering needs a period of low temperature induction (ie "vernalization") to start, and generally only blooms and bears fruit once a year. However, 'Sijimi' longan has the characteristic of continuous flowering (perpetual flowering, PF), which can continuously induce flowering and bear fruit in 1 year and 4 seasons. Dormant buds, flowers and fruits can exist on the same branch, and its flowering induction is not affected. External environmental influence. It is an excellent material for studying the analysis mechanism of longan flowering induction. In this study, three terminal buds at critical stages of flower induction were se...

Embodiment 2

[0038] Example 2 Subcellular Localization Analysis

[0039] Primers (terminator removal) were designed according to the cloned DlERF23 gene sequence (Table 1), and the full-length ORF of DlERF23 was amplified, and the PCR reaction procedure was as above. The PCR product was detected by 1% agarose gel electrophoresis, purified, connected to pMD 18-T carrier, and transformed into DH5α. Single colonies were picked, and plasmids were sequenced after PCR detection. Then the pBWA(V)HS-osgfp and DlERF23 plasmids were digested with EcoR I respectively, and enzyme ligation was carried out after recovery. The enzyme-linked plasmid was transformed into Escherichia coli DH5α, and after positive detection, the correct strain was selected for sequencing, and then extracted to obtain the pBWA(V)HS-DlERF23-osgfp plasmid. Then it was transferred into Arabidopsis protoplasts by PEG-mediated method (Yoo SD, Cho Y H, Sheen J.Arabidopsis mesophyllproto-plasts: a versatile cell system for transie...

Embodiment 3

[0040] Example 3 Overexpression vector construction and functional verification of transgenic Arabidopsis

[0041]Using specific PCR primers OEERF23-S / OEERF23-A (Table 1), longan cDNA was used as a template for PCR amplification. A BamH I restriction site is added to the 5' end of the primer, and a Sac I restriction site is added to the 5' end of the anti-primer. The obtained PCR product was ligated with pMD19-T vector and sequenced. Finally, the plasmids with correct sequencing were extracted, pBI121 and the plasmids with correct sequencing were double-digested with BamH I and Sac I, respectively, and a plant expression vector containing the DlERF23 target gene was constructed by T4 DNA ligase, and named pBI121-DlERF23. The constructed overexpression vector pBI121-DlERF23 was transformed into Agrobacterium strain GV3101 by liquid nitrogen freeze-thaw method, referring to the literature (Clough, Steven J, Bent, et al. Floral dip: simplified method for Agrobacterium-mediated t...

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Abstract

The invention provides a longan flowering regulation gene DlERF23 , its cDNA nucleotide sequence is shown in SEQ ID No.1. the longan DlERF23 The full length of the open reading frame of the gene is 552bp, encoding 183 amino acids, which has a typical AP2 / ERF domain and zinc finger structure, and belongs to the ERF protein. The results of qRT-PCR showed that the gene had tissue-specific expression, and the relative expression level was higher in pericarp, followed by stem, leaf and flower organs; the expression was down-regulated in the early flowering induction period of 'Sijimi' longan, while in the 'Sijimi' longan. The expression of Shixia'longan was up-regulated in the later stage of flowering. The results of transient expression in Arabidopsis protoplasts showed that the fluorescent signal was mainly concentrated in the nucleus. Transgenic Arabidopsis results showed that overexpression DlERF23 The transgenic plants showed early flowering: the wild-type plants flowered in about 26 days, while the transgenic lines flowered in 18‑20 days; the number of rosette leaves also increased. As a typical transcription factor, DlERF23 The encoded protein is located in the nucleus and positively regulates plant flowering, DlERF23 Gene overexpression significantly boosts plant flowering.

Description

technical field [0001] The invention relates to the field of molecular biology technology, in particular to a longan flowering regulation gene, its protein and its application. Background technique [0002] Flowering is an important event in the life cycle of plants, and the number and quality of flowers directly determine the final yield of plants. Floral transition is the process of plant transition from vegetative growth to reproductive growth, which is strictly controlled by endogenous developmental signals and external environmental factors, and is a complex signal "network". At present, the molecular genetic mechanism of flower induction mechanism in Arabidopsis and other model plants has been resolved. There are at least five major flowering pathways in Arabidopsis, including photoperiod, autonomous, vernalization, gibberellin, and aging pathways ( Turnbull C. Long-distance regulation of flowering time [J] Journal of Experimental Botany, 2011, 62(13): 4399-4413.). ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82C07K14/415C12Q1/6895A01H5/02A01H6/20
CPCC07K14/415C12N15/827C12Q1/6895C12Q2600/13C12Q2600/158
Inventor 决登伟桑雪莲石胜友唐建民
Owner 重庆圣盈达科技开发有限公司
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