Efficient, fast and stable gene transformation method for blueberries

A technology for gene transformation and blueberry, applied in the field of genetic engineering, can solve the problems of leaf browning and death, Agrobacterium and bacterial contamination, low blueberry transformation efficiency, etc., to prolong the infection time, reduce damage, and avoid Agrobacterium contamination. Effect

Active Publication Date: 2015-11-25
JIANGSU POLYTECHNIC COLLEGE OF AGRI & FORESTRY
View PDF3 Cites 6 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] We still face these problems that need to be solved: solve the pollution problem of Agrobacterium and miscellaneous bacteria; solve the problem of low transformation efficienc

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Efficient, fast and stable gene transformation method for blueberries
  • Efficient, fast and stable gene transformation method for blueberries

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1 The preparation of culture medium, bacterial strain and plant expression vector of the present invention

[0036] 1. Materials

[0037] (1) Plant material

[0038] The blueberry 'Meideng' tissue culture seedlings were subcultured on the blueberry subculture medium for experimentation.

[0039] (2) Preparation of main solution and culture medium

[0040] The medium preparation method is as follows:

[0041] Blueberry subculture medium (S1):

[0042] The improved WPM medium was supplemented with glucose 10g / L, phytogel 2.5g / L, ZT (zeatin) 1.5mg / L, NAA 0.4mg / L, pH 5.4, autoclaved, and set aside.

[0043] Blueberry co-cultivation medium (S2)

[0044] The improved WPM medium was supplemented with 30g / L sucrose, 5.5g / L agar, 6mg / L TDZ, pH 5.4, autoclaved, cooled to about 60°C and plated for later use.

[0045] Blueberry callus induction medium (S3)

[0046] Modified WPM medium supplemented with glucose 15g / L, plant gel 2.5g / L, ZT (zeatin) 1mg / L, TDZ1mg / L, ...

Embodiment 2

[0056] A method for efficient, rapid and stable gene transformation of blueberries, comprising the following steps:

[0057] 1) Strain activation: Streak the Agrobacterium AGL0 containing plasmid pK7WG2D on LB solid medium containing 50mg / L spectinomycin and 50mg / L rifampicin, and culture at 28°C for 2 days; Inoculate 2-3 single colonies from 50 mL of LB liquid medium containing 50 mg / L spectinomycin and 50 mg / L rifampicin, and culture in a shaker at 28°C and 220 rpm until the OD600 value is 0.4; at room temperature, Centrifuge at 5000rmp for 8 minutes, remove the supernatant, turn the centrifuge tube upside down on the sterile filter paper, remove the supernatant as much as possible, resuspend the bacteria with 100mL modified WPM liquid medium, and culture in a shaker for about 1 hour to measure OD600 The value is between 0.2 and 0.3;

[0058] 2) Plant material preparation: Take the young, flat 35-40-day-old leaves on the blueberry Mayden tissue culture seedlings, cut them a...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses an efficient, fast and stable gene transformation method for blueberries. The method comprises the following steps of 1 activation of culture, 2 plant material preparation, 3 preculture, 4 infection, 5 co-culture, 6 leaf washing, 7 callus induction, 8 bud induction and 9 root induction culture. According to the method, the gene transformation efficiency of the blueberries is improved, and the transformation rate can be above 10%. The concentration of bacterium liquid is reduced, and infection time is prolonged, so that agrobacterium pollution is avoided, and infection efficiency is improved. No drug or agentia assisting in infection is added to the infection bacterium liquid. Acetosyringone or other agentia assisting in infection is usually added in a traditional method, consequently, leaves are damaged, and the regeneration capacity of the leaves is reduced. Damage to the leaves of the blueberries is reduced, and the survival rate and regeneration rate of leaf culture are increased. A carrier carrying high expression and reporter genes is adopted, transgenic plants can be directly screened out through observation, and tedious work in traditional detection is avoided. The period of transgenosis is shortened.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, in particular to a method for efficient, rapid and stable gene transformation of blueberries. Background technique [0002] Blueberry (Vaccinium spp.) is a new kind of fruit tree, which has been widely cultivated in many countries in recent years, and is one of the excellent fruit trees with very rapid development. British authoritative nutritionist lists blueberry as the first of 15 kinds of healthy foods, and the Food and Agriculture Organization of the United Nations also lists blueberries as one of the top five healthy foods for human beings. As a kind of fruit tree rich in nutrition, with great health care function and high economic value, blueberry will surely receive more and more attention today when people's living standards are constantly improving. With the continuous development of science and the continuous innovation of technology, the research on blueberry will gradual...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N15/82A01H5/00
Inventor 王媛花冯英娜史红林
Owner JIANGSU POLYTECHNIC COLLEGE OF AGRI & FORESTRY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap