Micropropagation by Somatic Embryogenesis of Coffea Species and Cryopreservation

咖啡物种体细胞胚胎发生的微繁殖和冷冻保存

• 批准号: 04660015
• 负责人: YASUDA Takeshi
• 金额: $ 1.28万
• 依托单位: Kobe University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04660015/
• 关键词: Somatic embryo; Coffee; Cryopreservation; Cytokinin; Protoplast; Tropical Tree; Coffea; 凍結保存

The embryogenic callus and somatic embryo of Coffea arabica and C.canephora can be induced by cytokinin alone from the section of young leaf of mature trees. The cytokinin induced callus has a high embryogenic potetial and is very useful for clonal propagation. As cytokinin, 2-isopentenyl-adenine(2iP)is the best, and benzyladenin(BA), zeatin and kinetin also are effective for somatic embryogenesis from coffee leaf slices. Additional auxins act inhibitory effect on somatic embryogenesis. Protoplast were prepared from embryogenic calluses from C.arabica and canephora and cultured on Gelrite medium with cytokinin as a plant regulator. Somatic embryos were regenerated from protoplasts. These simplified method for preparation and culture of protoplasts is very useful for cell fusion and genetic engeering. Somatic embryos of Coffea were successfully cryopreserved after being exposed to a series of preparative steps ; precultured on media with increasing concentration of sucrose, encapsulated in alginate beads, dehydrated up to 30% and pluged into liquid nitrogen.

用细胞分裂素单独诱导小粒咖啡和中粒咖啡成熟树幼叶的胚性愈伤组织和体细胞胚。细胞分裂素诱导的愈伤组织具有很高的胚性发生潜力，对无性繁殖非常有用。作为细胞分裂素，2-异戊烯基腺嘌呤（2 iP）效果最好，苄基腺嘌呤（BA）、玉米素和激动素对咖啡叶片体细胞胚胎发生也有一定的促进作用。附加生长素对体细胞胚胎发生有抑制作用。从小粒种和中杯种的胚性愈伤组织中制备原生质体，并将其培养在含有细胞分裂素的Gelrite培养基上。原生质体再生体细胞胚。这些简化的原生质体制备和培养方法对细胞融合和遗传工程具有重要意义。在暴露于一系列制备步骤之后，成功地冷冻保存了咖啡属的体细胞胚;在具有增加浓度的蔗糖的培养基上预培养，包封在藻酸盐珠中，脱水至30%并插入液氮中。

项目成果

TAHARA: "Formation of somatic embryos from protoplasts of Coffea arabica" Hort Science. 29(印刷中). (1994)

田原：“阿拉比卡咖啡原生质体的形成”Hort Science 29（出版中）。

山口 康之: "植物分子・細胞工学マニュアル" 講談社サイエンティフィック, 228 (1992)

山口康之：《植物分子和细胞工程手册》讲谈社科学，228（1992）

安田武司: "組織培養のepoch-making2.コーヒーの組織培養" 植物細胞工学. 5. 53-57 (1993)

安田武：“组织培养划时代2.咖啡组织培养”植物细胞工程5. 53-57 (1993)。

HATANAKA: "Direct regrowth of encapsulated somatic embryos of coffee(coffea canephora)after cooling in liquid nitrogen" Cryo-Letters. 15. 47-52 (1994)

HATANAKA：“咖啡（coffea canephora）的封装体细胞胚胎在液氮中冷却后直接再生”Cryo-Letters。

HATANAKA: "Direct regrowth of encapsulated somatic embryos of coffee(Coffea canephora) after cooling in liquid nitrogen" Cryo-Letters. 15. 47-52 (1994)

HATANAKA：“咖啡（Coffea canephora）的封装体细胞胚胎在液氮中冷却后直接再生”Cryo-Letters。