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Phoebe bournei somatic embryo and adventitious bud induction method

A technology of somatic embryos and cell embryos, which is applied in the field of induction and cultivation of somatic embryos in Minnan, can solve the problems of prominent seed size and age, no research reports on rapid propagation technology, and few resources in Minnan, so as to improve the quality of zygotic embryos. Effects of somatic embryo induction rate, expansion of adventitious bud reproduction coefficient, and improvement of emergence rate

Active Publication Date: 2017-06-20
ZHEJIANG FORESTRY UNIVERSITY
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AI Technical Summary

Problems solved by technology

[0003] The natural reproduction of Minnan is mainly through seeds, but the size of the seeds is prominent, and the seeds have a short lifespan in the wild, and the germination rate is extremely low, which leads to very few resources of Minnan.
In recent years, a large number of scientific researchers have begun to study related technologies of Nannan seedling conservation, but most of them focus on the vitality, dormancy and germination of Nannan seeds (Li Tiehua, Wen Shizhi, Peng Xianfeng, etc., 2003, 2009, 2010), seedling cultivation (Liu Jun, Jiang Jingmin, Chen Yitai, etc., 2011), although tissue culture techniques have also been reported (Qu Fenxia and Chen Cunji, 2010), but their proliferation rate is low, and the use of modern biotechnology to carry out the study of Minnan There is no research report on the rapid propagation technology of embryogenic callus and somatic embryo induction culture

Method used

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  • Phoebe bournei somatic embryo and adventitious bud induction method
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  • Phoebe bournei somatic embryo and adventitious bud induction method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 Screening of Somatic Embryo Induction Medium and Embryogenic Callus Induction Medium of Minnan Seed Immature Embryos

[0023] The immature Minnan seeds are used as materials. Soak in detergent for 1 minute, rinse with tap water for 3 hours, rinse with sterile water twice, treat with 75% alcohol for 30 seconds, rinse with sterile water for 3 to 4 times, disinfect with 20% (volume ratio) NaClO for 15 minutes, rinse with sterile water for 3 to 4 times , took out immature embryos under aseptic conditions, pretreated in 0.5M sucrose at 4°C for 48 hours, and inoculated in 13 solid medium containing different hormone combinations of MS+hydrolyzed casein 500mg / L+L-glutamine 500mg / L for in vivo Cell embryogenesis experiments. Each treatment was inoculated into 3 petri dishes (specification: 90*15mm), and 5 immature embryos were inoculated in each dish, 15 inoculations in total, and 4 replicates. The relative humidity of the culture room is 70%±5%, the temperature is ...

Embodiment 2

[0032] Example 2 Effects of Different Young Embryo Development Stages on Somatic Embryo Induction Rate and Healing Rate

[0033] On August 5th, August 12th, August 19th, August 26th, September 3rd, and September 10th, 2016, respectively, the seeds of strong and sunny branches of excellent plants were collected, and one excellent plant was used as a repetition. The 3 superior plants were repeated 3 times, and each time 75 seeds were collected from each superior plant, 30 seeds were used for callus induction, 30 seeds were used for somatic embryo induction, and 15 seeds were used for dissection observation to determine the zygote in each time period stage of embryonic development. The explant pretreatment method and culture environment are the same as in Example 1. The medium for somatic embryo induction is MS+hydrolyzed casein 500mg / L+L-glutamine 500mg / L+2,4-D 1.5mg / L+6-BA 1.0mg / L, and the medium for callus induction is MS+ Hydrolyzed casein 500mg / L+L-glutamine 500mg / L+IBA 1....

Embodiment 3

[0037] Example 3 Somatic Embryo Proliferation and Culture

[0038] The obtained primary embryos were transferred to the proliferation medium for proliferation and culture to produce secondary embryos. The medium used MS as the basic medium. The combination of hormone addition and proliferation results are shown in Table 3. The experimental design and culture conditions are the same as in Example 1. Observe daily after inoculation, and count the results after 20 days.

[0039] Table 3 Effects of different proliferation media on the proliferation of somatic embryos

[0040] 1 2 3 4 5 6 6-BA 0.5 0.5 0.5 1.0 1.0 1.0 NAA 0.1 0.5 1.0 0.1 0.5 1.0 Multiplication coefficient 5.00 12.9 6.5 6.2 4.7 3.0

[0041] Note: Proliferation coefficient = number of secondary embryos produced / number of primary embryos inoculated

[0042] One week after the inoculation, it was found that some of the primary embryos had a white embryo-like structu...

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Abstract

The invention discloses a phoebe bournei somatic embryo and adventitious bud induction method. The phoebe bournei somatic embryo and adventitious bud induction method comprises the following technical procedures: by taking an immature seed of phoebe bournei as a material, stripping a young embryo of the immature seed as an explant, carrying out somatic embryo induction and enrichment culture to generate a secondary embryo, or carrying out embryonic callus induction, differentiating adventitious buds and the like. According to the phoebe bournei somatic embryo and adventitious bud induction method, a somatic embryo and embryonic callus induction and culture technical system for an immature zygotic embryo of the phoebe bournei is built for the first time, so that the optimal time for taking materials is specified, and an excellent somatic embryo, an embryonic callus induction culture medium, an enrichment culture medium, a somatic embryo germination culture medium and a differentiation culture medium are screened out; a somatic embryo induction rate, an enrichment rate, a somatic embryo germination rate, a differentiation coefficient and the like are effectively increased, and the current situation of severe short supply of phoebe bournei seeds can be relieved. Meanwhile, successful induction of embryonic callus provides a technical guarantee for somatic fusion, preservation of seed resources, gene transformation and excellent hybridization and combination for propagation expansion.

Description

technical field [0001] The invention belongs to the field of plant biotechnology, and in particular relates to a method for inducing and cultivating somatic embryos of Phoebe fraternis. Background technique [0002] Minnan (Phoebe bournei (Hemsl.) Yang), endemic to China, is a national second-class rare and endangered species (Fu Liguo, Jin Jianming. Chinese Plant Red Book of Rare and Endangered Plants, Science Press, 1991). Minnan is a large evergreen tree of Lauraceae, distributed in Jiangxi, Fujian, southern Zhejiang, Guangdong, Guangxi and other places in the mid-subtropical evergreen broad-leaved forest zone (Chinese Academy of Sciences Chinese Flora Editorial Committee Chinese Flora: Volume 31, Beijing : Science Press, 1982.). Minnan wood is fragrant and durable, light yellow, dense and tough material, not easy to warp and crack, easy to process, smooth beveled surface, beautiful texture, it is a good wood for high-grade buildings, furniture, shipbuilding and craft ca...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/005A01H4/008
Inventor 童再康楼雄珍黄华宏张俊红林二培
Owner ZHEJIANG FORESTRY UNIVERSITY
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