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Inducing method for adventitious buds of mulberry cotyledons

A bud induction and mulberry technology, applied in the field of plant genetic engineering, can solve the problems of low cytokinin and auxin concentrations, lack of histology and anatomy, low cotyledon adventitious bud induction rate, etc., and achieve stable results and induction effects. Good, short time period effect

Inactive Publication Date: 2014-09-10
JIANGSU UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Scholars such as Wang Yong disclosed the induction and regeneration of mulberry leaf adventitious buds, using mulberry leaves as recipients to prepare for mulberry genetic engineering research, but the concentration of cytokinin and auxin is relatively low, and the method of induction is adopted once , regardless of the different hormone concentration requirements for adventitious bud formation and development, so the induction rate of cotyledon adventitious buds is very low; in addition, this method lacks histological and anatomical methods to prove whether the induced buds are from adventitious buds or embryos

Method used

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  • Inducing method for adventitious buds of mulberry cotyledons
  • Inducing method for adventitious buds of mulberry cotyledons
  • Inducing method for adventitious buds of mulberry cotyledons

Examples

Experimental program
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Effect test

Embodiment 1

[0023] (1) Seed imbibition and cultivation

[0024] Take 200 first-generation hybrid Sangfengchi mulberry seeds, put them in a beaker and soak in water for 12 hours, then put the beaker under the tap and rinse it with running water for 2 hours; then drain the water in the beaker, add 75% alcohol to shake for 10 minutes, pour Rinse twice with clean water after drying alcohol; add 7% sodium hypochlorite solution to sterilize the surface for 15 minutes, shake continuously during this period, move it into an ultra-clean workbench, and rinse with sterilized distilled water 3 times.

[0025] (2) Separation of cotyledons

[0026] Put the surface-sterilized seeds on the dissecting mirror in the ultra-clean workbench, use a scalpel to cut the seed coat gap on the side of the hilum, use tweezers to lightly press the opposite side of the seed coat gap, and gently squeeze out the complete embryo Gently separate the two cotyledons in the embryo with dissecting forceps, and cut off the cot...

Embodiment 2

[0037] (1) Seed imbibition and cultivation

[0038] Take 200 mulberry seeds, put them in a beaker and add water to soak for 12 hours, then put the beaker under the tap and rinse it with running water for 2 hours; then drain the water in the beaker, add 0.1% mercuric chloride solution to sterilize the surface for 15 minutes, and keep Shake it, move it into the ultra-clean workbench, pour out the mercuric chloride solution, and rinse it with sterilized water for 3 times.

[0039] Put the sterilized seeds on the surface into the germination medium (1 / 2MS medium + 15g / L sucrose + 7g / L agar powder, pH value 6.0), and cultivate them for two days before separating the cotyledons.

[0040] (2) Separation of cotyledons

[0041] Place the cultivated seeds on the dissecting mirror in the ultra-clean workbench, use a scalpel to incise the seed coat gap on the side of the hilum, use two tweezers to gently tear the seed coat gap, and completely peel off the embryo. The two cotyledons in t...

Embodiment 3

[0051] (1) Seed imbibition and cultivation

[0052] Take 200 Guangdong hybrid mulberry seeds, put them into a beaker and add a little water to soak for 12 hours, then put the beaker under the tap and rinse it with running water for 2 hours. Drain the water in the beaker, add 0.1% mercuric chloride solution to sterilize the surface for 15 minutes, shake it constantly, move it into the ultra-clean workbench, pour out the mercuric chloride solution, and rinse it with sterilized water three times.

[0053] Put the sterilized seeds on the surface into the germination medium (1 / 2MS medium + 15g / L sucrose + 7g / L agar powder, pH value 6.0), during which there are seeds contaminated by bacteria, and move the uncontaminated seeds into a new germinated medium medium.

[0054] (2) Separation of cotyledons

[0055] The seeds cultivated for 3 days were placed on the internal dissecting mirror of the ultra-clean workbench, and the seed coat gap was cut on the side of the hilum with a scalp...

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Abstract

The invention discloses an inducing method for adventitious buds of mulberry cotyledons, and belongs to the field of plant gene engineering. The inducing method comprises the steps of with cotyledons in mulberry seed entoderm as explants, performing surface sterilization on seeds washed by running water and soaked, performing separation to obtain the cotyledons, then directly inducing the cotyledons in an inducing culture medium to generate adventitious buds, and transferring the adventitious buds into a rooting culture medium when the adventitious buds are 1cm long, thereby forming complete plants. According to the method, regenerated plants produced by the adventitious buds are short in time period, results are stable, and the repetitiveness is high; the inducing method is mainly used for mulberry transgenosis and supplies a premise and a basis for mulberry transgenosis breeding.

Description

technical field [0001] The invention relates to a method for directly inducing cotyledon adventitious buds, which is a transgene acceptor part of mulberry trees and belongs to the field of plant genetic engineering. Background technique [0002] Among many plant regeneration systems, the method of inducing leaf adventitious buds, somatic embryos and protoplast regeneration belongs to the single-cell origin, which is suitable for the receptor system of plant transgenesis. Among many mature transgenic technologies, plants such as peanuts and poplars mostly use the method of infecting leaves to produce adventitious buds to regenerate plants, and crops such as cotton and rice mostly use the method of infecting hypocotyls to induce embryogenic callus and regenerate plants through somatic embryos. Or induce embryogenic callus first, then infect embryogenic callus, and regenerate plants through somatic embryo germination. The protoplast system is rarely used as a transgenic recept...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
Inventor 潘刚邸炳荣胡颖健孙银苹雷朋
Owner JIANGSU UNIV OF SCI & TECH
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