Somatic embryogenesis from mature split seeds of jaboticaba (Plinia peruviana (Poir) Govaerts)

ABSTRACT. Plinia peruviana is a species that is native to Brazil and is important due to the taste and medicinal properties of its fruits. Young leaves and split mature seeds were used as explants to initiate somatic embryogenesis to obtain a large number of plants in a short period of time. Leaf discs were cultured in MS medium containing various concentrations of 2,4-D (2,4-dichlorophenoxyacetic acid) or picloram (4-amino-3,5,6-trichloro-2-pyridinecarboxylic acid). In the case of the mature seeds, various concentrations of glutamine, 2,4-D and a combination of auxin and BAP (6-benzylaminopurine) were tested for somatic embryogenesis induction. For somatic embryo maturation, several concentrations of PEG 6000 (polyethylene glycol; up to 90 g L-1) were tested. After 60 days of culture using leaf discs, callus formation occurred in all treatments, with the highest averages obtained with 10 μM 2,4-D. However, these calluses did not form somatic embryos. For the cultured seeds, the best treatment was the MS medium with 1,000 mg L-1 glutamine and 10 μM 2,4-D without BAP. The supplementation of 60 g L-1 PEG 6000 was sufficient to promote the maturation of the somatic embryos. Histological analyses of the calluses that were formed from leaf discs showed nonembryogenic characteristics. In contrast, the calluses that originated from mature seeds had small and round cells with little vacuolation, which are characteristics of embryogenic structures.

摘要：Plinia peruviana为原产于巴西的物种，其果实兼具优良风味与药用特性，因此具有重要应用价值。本研究以幼叶与裂瓣成熟种子作为外植体（explant），诱导体细胞胚胎发生（somatic embryogenesis），以期在短时间内批量获得再生植株。将叶盘接种于添加不同浓度2,4-二氯苯氧乙酸（2,4-dichlorophenoxyacetic acid，简称2,4-D）或氯氨吡啶酸（picloram，4-氨基-3,5,6-三氯-2-吡啶羧酸）的MS培养基中培养；针对成熟种子组，本研究测试了不同浓度谷氨酰胺、2,4-D以及生长素与6-苄氨基腺嘌呤（6-benzylaminopurine，简称BAP）的组合对体细胞胚胎发生的诱导效果。在体细胞胚成熟阶段，本研究测试了不同浓度的聚乙二醇6000（polyethylene glycol 6000，简称PEG 6000；最高浓度为90 g·L⁻¹）的作用效果。叶盘培养60天后，所有处理组均诱导产生愈伤组织（callus），其中10 μM 2,4-D处理组的愈伤组织诱导率均值最高，但此类愈伤组织均未形成体细胞胚。成熟种子组的最优处理为添加1000 mg·L⁻¹谷氨酰胺与10 μM 2,4-D且不含BAP的MS培养基。添加60 g·L⁻¹ PEG 6000即可有效促进体细胞胚的成熟。对叶盘来源的愈伤组织进行组织学分析后发现，其具备非胚性结构特征；与之相反，成熟种子来源的愈伤组织由体积小巧、形态圆润且液泡化程度低的细胞构成，这正是胚性结构的典型特征。