A kind of breeding method of using ethyl methanesulfonate to induce in vitro mutagenesis of Ciba japonica

A kind of ethyl methanesulfonate, consistent technology, applied in the breeding field of using ethyl methanesulfonate to induce in vitro mutagenesis of Cizhu liangshanensis, can solve the problems such as unseen mutation breeding

Active Publication Date: 2021-08-06
SOUTHWEAT UNIV OF SCI & TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, there have been no research reports on mutation breeding in bamboo plants

Method used

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  • A kind of breeding method of using ethyl methanesulfonate to induce in vitro mutagenesis of Ciba japonica
  • A kind of breeding method of using ethyl methanesulfonate to induce in vitro mutagenesis of Ciba japonica
  • A kind of breeding method of using ethyl methanesulfonate to induce in vitro mutagenesis of Ciba japonica

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Embodiment 1, preparation bamboo callus

[0069] 1. Preparation of explants

[0070] 1. Take the young side branches of Cizhu liangshan, remove the leaf sheaths, and then carry out the following disinfection steps in sequence: wash with 70% ethanol aqueous solution, then rinse with running water, then soak for 30 sec with 70% ethanol aqueous solution, then fully clean with sterile water, Then with 0.1% HgCl 2 The aqueous solution was disinfected for 10 minutes, and then fully rinsed with sterile water.

[0071] 2. After completing step 1, take side branches and cut them into stem segments with a length of 1±0.2cm (each stem segment has at least one node).

[0072] 2. Preparation of callus (the culture temperature in the whole process is 25±2°C)

[0073] 1. Take the stem segment obtained in step 1, inoculate it on the callus induction medium, cultivate it for 2 weeks (dark culture), and then cultivate it for 6-8 weeks (16 hours of light per day) to obtain callus.

[...

Embodiment 2

[0079] Embodiment 2, utilization of ethyl methanesulfonate in vitro mutagenesis of Cizhu liangshan

[0080] 1. Prepare pH 4.8, 0.01M phosphate buffer, autoclave and cool to room temperature.

[0081] 2. Take ethyl methanesulfonate, filter and sterilize with a 0.22 μm microporous membrane.

[0082] 3. Take the ethyl methanesulfonate obtained in step 2 and dilute it with the phosphate buffer obtained in step 1 to obtain an ethyl methanesulfonate solution.

[0083] In the ethyl methanesulfonate solution, the volume percent content of ethyl methanesulfonate is 0.6%, 0.8% or 1.0%, respectively.

[0084] 4. Take the callus obtained in Example 1 (the number is n1), place it in ethyl methanesulfonate solution, and shake it for 30 minutes.

[0085] 5. After completing step 4, take the callus and rinse it with sterile water.

[0086] 6. After step 5 is completed, the callus is taken, inoculated into the cluster bud differentiation medium, and cultured until the cluster shoots grow ou...

Embodiment 3

[0104] Example 3. Analysis of Mutagenized Progeny Using ISSR Molecular Marker Technology

[0105] From the 96 regenerated plants obtained after mutagenesis with 0.6% ethyl methanesulfonate solution in Example 2, 47 were randomly selected as candidate mutants.

[0106] Each candidate mutant strain and control strain were analyzed by ISSR molecular marker technology.

[0107] 1. Take leaves and extract genomic DNA.

[0108] 2. Using the genomic DNA obtained in step 1 as a template, carry out ISSR-PCR amplification with ISSR primers.

[0109] The 14 ISSR primers in Table 2 were used respectively. In Table 2, Y stands for C or T, R stands for A or G, B stands for C or G or T, D stands for A or G or T, and S stands for G or C.

[0110] Table 2

[0111] Primer name Primer sequence (5'-3') ISSR-A AGAGAGAGAGAGAGAGYA ISSR-B GAGAGAGAGAGAGAGAYC ISSR-C (Sequence 1 of the Sequence Listing) CTCTCTCTCTCTCTCTCTRC ISSR-D (Sequence 2 of the Sequence Lis...

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Abstract

The invention discloses a breeding method for using ethyl methanesulfonate to induce in vitro mutagenesis of Cizhu japonica. The invention provides a method for obtaining mutant plants of Arthia lanceolata, comprising the following steps: using ethyl methanesulfonate to mutate the callus tissue derived from Arthia lanceolata. The method for obtaining the mutant plant of Cizhu liangshanensis also includes the following steps: cultivating the callus tissue after mutagenesis as a regenerated plant. The method for obtaining the mutant plants of Cizhu liangshanensis further comprises the following steps: screening the mutant plants from the regenerated plants. In the method provided by the invention, the mutagenesis technique is combined with the in vitro culture, and ethyl methanesulfonate is used to in vitro mutagenize the callus of Cizhu liangshanensis, so that a large number and homogeneous mutagenized population can be obtained. The invention greatly improves the mutation frequency and the breeding efficiency, is beneficial to improving the mutation breeding of the Liangshan Cizhu, will promote the industrial development of the Liangshan Cizhu, and has great application and popularization value.

Description

technical field [0001] The invention belongs to the field of plant mutation breeding, and in particular relates to a breeding method for using ethyl methanesulfonate to induce mutagenesis of Ciba japonica in vitro. Background technique [0002] The pulp and paper industry currently has the problem of relatively low recycling rate of waste paper. Bamboo, as a wide range of plant natural resources, has the characteristics of wide distribution, strong adaptability, fast growth, early maturity, and high economic value. After a successful afforestation, it can be felled every year after 3-5 years, and lasts for decades. Even hundreds of years. The cellulose content of bamboo is 40%-60%, its excellent pulping performance can be compared with some wood fibers, and the fiber shape quality of bamboo pulp is between straw pulp and wood pulp, closer to wood pulp. Prompting it to become an excellent non-wood papermaking raw material. Therefore, cultivating bamboo varieties with large...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 胡尚连曹颖黄艳龙治坚徐刚任鹏卢学琴赵博
Owner SOUTHWEAT UNIV OF SCI & TECH
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