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Transcription factor EjBZR1 for inhibiting fruit cell expansion and application thereof

A technology of transcription factors and cells, applied in application, genetic engineering, plant genetic improvement, etc., can solve problems such as failure to find

Active Publication Date: 2020-02-11
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the key genes that regulate the size of loquat fruit have not been found

Method used

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  • Transcription factor EjBZR1 for inhibiting fruit cell expansion and application thereof
  • Transcription factor EjBZR1 for inhibiting fruit cell expansion and application thereof
  • Transcription factor EjBZR1 for inhibiting fruit cell expansion and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Cloning of embodiment 1 loquat EjBZR1 gene cDNA sequence

[0049] 1. Experimental method

[0050] 1. Sample collection

[0051] The four developmental stage fruit samples of ‘Zaozhong 6’ were collected from the production trees of ‘Zaozhong 6’ in full fruit stage planted in the loquat germplasm resource nursery of South China Agricultural University. Fresh fruit samples were quickly frozen in liquid nitrogen and stored in a -80°C freezer.

[0052] 2. Extraction of RNA

[0053] Use the EASYspin Plus Plant RNA Rapid Extraction Kit (Adelaide) to extract RNA from the ground fruit samples according to the instructions, as follows: Take 1.0mL RLT lysate and add it to a 1.5mL centrifuge tube, add 100mg of ground fruit sample powder , shake vigorously for 20s, then lyse at 55°C for 20min; centrifuge the lysate at 13,000rpm for 5-10min; take the supernatant, transfer it to a new centrifuge tube, add absolute ethanol half the volume of the supernatant, and gently pipette to mi...

Embodiment 2

[0065] Example 2 Analysis of the expression pattern of EjBZR1 during the growth of ZP44 and ZP65 fruits

[0066] 1. Experimental method

[0067] Loquat fruit samples were collected at different stages of loquat fruit development and fruit / receptacle diameter, thickness and weight were determined. Fruit samples of 'ZP44' and 'ZP65' were collected at 10 different periods after fruit setting for comparison of fruit weight, cell characteristics and gene expression levels. All fruit measurements were taken in 3 biological replicates, and 10 fruits were taken for each replicate.

[0068] Table 1 gene expression analysis primer pair sequence list

[0069]

[0070] Make slices, as follows: Use FAA fixative to fix the small pieces of fruit pulp in each period, and place them in a vacuum pump for about 1 hour to filter until the small pieces of pulp floating in the fixative sink to the bottom of the bottle to ensure that the cells are fully fixed; Take out Put the sample block into...

Embodiment 3

[0074] Example 3 Construction of EjBZR1 gene silencing vector and VIGS treatment of loquat fruit

[0075] 1. Experimental method

[0076]EjBZR1 was subcloned into the TRV2 vector using the primers in Table 2 to construct the virus-induced gene silencing vector TRV2-EjBZR1. The carrier construction is specifically: using Gibson Master Mix and corresponding endonucleases (purchased from NewEngland Bio labs) were used for the linearized vector at 37°C. The digested products were purified using Gel DNA Mini Recovery Kit (Magen). Use the ClonExpress one-step directional cloning seamless cloning kit to connect the purified enzyme-digested product to the target vector. The reaction system contains 1 μl purified and recovered PCR amplification product, 1 μl linearized vector, 2 μl 5×CE II buffer, 1 μl Exnase TM II and 4 μl sterile water. The reaction system was reacted at 37°C for half an hour, then cooled on ice for 5 minutes, and then 5 μl of the ligation product was transferre...

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Abstract

The invention discloses a transcription factor EjBZR1 for inhibiting fruit cell expansion and application thereof. The BES1 / BZR1 family member EjBZR1 gene for regulating cell expansion and fruit growth in loquat fruits is isolated and cloned from ' Zaozhong No. 6 '. The gene has a nucleotide sequence shown as SEQ ID No. 1 and comprises an open reading frame of 888bp; and encodes 295 amino acids, and the encoded amino acid sequence is shown in SEQ ID No. 2. The transcription factor gene provides a gene target for regulating cell size and the fruit size of loquat fruits, provides a theoretical basis for improving sizes of product organs such as fruits and other crop grains, and ensuring crop yield, provides a new molecular breeding gene resource for reducing use of growth regulators in fruitproduction, and provides a new crop genetic resource for production of environment-friendly agricultural products.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, more specifically, to a transcription factor EjBZR1 that inhibits fruit cell expansion and its application Background technique [0002] Loquat, which sprouts winter flowers in autumn and ripens in summer in spring, is a characteristic and precious fruit native to my country. The yield per mu of ordinary loquat orchards is about 500kg, but only a few well-managed orchards can reach 1000kg per mu, which is less than 30% of that of tree species such as apples and citrus. At present, the high production cost caused by low yield per unit area has become an important resistance to the development of loquat industry. Fruit size is not only an important quality trait of fruit, but also an important basis for yield formation. There are a large number of single flowers on each inflorescence of loquat. Although each ear can naturally set 10 to 20 fruits, the growth of these fruits is un...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/82C07K14/415A01H5/00A01H6/20A01H6/74
CPCC07K14/415C12N15/8218C12N15/8261
Inventor 苏文炳林顺权杨向晖邵子坤甘小清
Owner SOUTH CHINA AGRI UNIV
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