Rose skin thorn regulation gene RrCPC and application thereof

A technique for regulating genes, roses

Active Publication Date: 2022-05-06
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, rose stems are thorny, and there are many inconveniences in the process of garden application, field management and flower picking. If rose varieties with few or no thorns can be cultivated, the work efficiency of rose field management and picking will be greatly improved. Improving the Economic Benefits of Rose Planting

Method used

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  • Rose skin thorn regulation gene RrCPC and application thereof
  • Rose skin thorn regulation gene RrCPC and application thereof
  • Rose skin thorn regulation gene RrCPC and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] 1. Clone by RACE technology RrCPC Gene;

[0021] Based on rose transcripts, RACE primers were designed to use the total RNA of rose organs as a template to obtain 5' and 3' end fragments, clone them into T-vectors, and perform sequencing after positive screening of the insert fragments. The sequences of the sequencing results were spliced ​​through overlapping regions , to obtain the full-length cDNA.

[0022] I. Primer design:

[0023] 3'RACE-specific primers:

[0024] Outer Primer: 5'-GAAGAGGTAAGCAGCATT-3';

[0025] Inner Primer: 5'-CAGGTCGGAAAGCAGAAG-3';

[0026] 3'RACE Adapter Primer:

[0027] Outer Primer: 5'-TACCGTCGTTCCACTAGTGATTT-3';

[0028] Inner Primer: 5'-CGCGGATCCTCCACTAGTGATTTCACTATAGG-3';

[0029] CGCAGAGT-3' was used for the universal linker primer.

[0030] 5'RACE Adapter Primer:

[0031] Outer Primer: 5'-CTAATACGACTCACTATAGGGCAAGCAGTGGTATCAACGCAGAGT-3';

[0032] Inner Primer: 5'-CTAATACGACTCACTATAGGGC-3';

[0033] 5'RACE-specific primers:

...

Embodiment 2

[0070] The temporal and spatial expression pattern of RrCPC gene was explored by fluorescent quantitative PCR technology, and fluorescent quantitative PCR primers were designed based on the exon region of RrCPC gene. The internal reference primers were designed as forward primer 5'-TGAGGCCATTTACGACAT-3' and reverse primer 5'-AGATCACAGGAGCATAGGAG-3'. Utilize SYBR Premix ExTaq (Takara Company) and fluorescent quantitative PCR instrument CFX96TM (Bio-RAD Company), refer to the PCR system and program of manufacturer's instructions, carry out fluorescent quantitative PCR to obtain the number of cycles reaching the fluorescence threshold, calculate the RrCPC gene in purple branch The relative expression levels on the first, second and third lateral branches of rose.

[0071] Such as figure 1 As shown, the expression level of RrCPC gene was the lowest in the epidermis tissue of the first collateral branch (FCB), slightly increased in the epidermal tissue of the secondary collateral...

Embodiment 3

[0072] Example 3 RrCPC Overexpression Vector Construction and Arabidopsis Genetic Transformation

[0073] In order to better elucidate the function of the RrCPC gene, the applicant overexpressed it in Arabidopsis plants, and performed functional verification according to the phenotype of the transgenic plants. The method is as follows: insert the RrCPC expression cassette into the multiple cloning site of the overexpression vector pCAMBIA1304 using the NcoI restriction site ( figure 2 ), the obtained plasmid was transformed into Agrobacterium EHA105 competent by liquid nitrogen freeze-thaw method, and pCAMBIA1304: RrCPC Agrobacterium positive strain was identified.

[0074] Infect Arabidopsis thaliana by inflorescence dipping method (conventional method), and obtain transgenic plants after screening transformed seedlings with hygromycin resistance.

[0075] The main steps and application reagents of the genetic transformation of the present invention are as follows:

[007...

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Abstract

The invention relates to a rose skin thorn regulation and control gene RrCPC and application thereof, and the RrCPC gene is cloned by taking purple-branch rose as a material through RACE (Rapid Amplification of cDNA Ends) technology. The expression mode of the RrCPC gene on different levels of lateral branches of roses is detected through a real-time fluorescent quantitative detection technology, meanwhile, arabidopsis thaliana is transformed by constructing an overexpression vector, it is proved that the RrCPC gene is a negative regulatory factor for epidermal hair generation, and the RrCPC gene has important application value for cultivating few-thorn or thorn-free rose new germplasm and improving rose management efficiency. The invention provides a rose skin thorn regulation gene RrCPC and application thereof in few-thorn and thorn-free roses, and the rose skin thorn regulation gene RrCPC has important application value in the field of thorn-free rose breeding.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, in particular to a rose thorn regulation gene RrCPC and its application. Background technique [0002] Rose( Rosa rugosa Thunb . ) is a deciduous shrub belonging to Rosaceae Rosaceae, with pleasant fragrance and various varieties. It is a traditional famous flower in my country, a precious natural spice plant and an excellent landscaping material. Its flowers can extract rose essential oil and make rose tea, rose sauce, etc., which are widely used in spices, food, cosmetics and other industries, with high economic and health value. [0003] With the development of social economy and the improvement of people's living standards, the demand for rose products in the international and domestic markets is gradually increasing, and it is showing a variety of characteristics. The requirements for the aroma, quality, ornamental and variety of roses are also increasing. However, ros...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82A01H5/00A01H6/74
CPCC07K14/415C12N15/8212C12N15/8226C12N15/8261Y02A50/30
Inventor 冯立国储亚东王建文徐勇魏国
Owner YANGZHOU UNIV
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