Ouyang Yao, Chen Yulu, Lü Jinfeng, Teixeira da Silva Jaime A, Zhang Xinhua, Ma Guohua
Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, Guangzhou, 510650, China.
University of Chinese Academy of Sciences, Beijing 100039, China.
Sci Rep. 2016 Apr 19;6:24662. doi: 10.1038/srep24662.
An efficient protocol providing a dual regeneration pathway via direct shoot organogenesis and somatic embryogenesis for an endangered species, Metabriggsia ovalifolia W. T. Wang, was established from leaf explants. When applied at 2.5 μM, the cytokinins 6-benzyladenine (BA) or thidiazuron (TDZ) and the auxins indole-3-butyric acid (IBA), α-naphthaleneacetic acid (NAA) and indole-3-acetic acid (IAA) could induce shoots when on basal Murashige and Skoog (MS) medium. BA and TDZ could induce more adventitious shoots (19.1 and 31.2/explant, respectively) than NAA (4.6/explant), IBA (5.7/explant) or IAA (6.4/explant). BA and TDZ at 5-10 μM could induce both shoots and somatic embryos. A higher concentration of TDZ (25 μM) induced only somatic embryos (39.8/explant). The same concentration of BA induced both adventitious shoots (23.6/explant) and somatic embryos (9.7/explant). Thus, somatic embryogenesis in this plant needs a high cytokinin concentration (BA; TDZ), as evidenced by histology. Somatic embryos germinated easily when left on the same media, but formed adventitious roots in two weeks on MS supplemented with 0.5 μM NAA, 0.5 μM IBA and 0.1% activated charcoal. Over 93% of plantlets survived following acclimatization and transfer to a mixture of sand and vermiculite (1:1, v/v) in trays.
通过直接芽器官发生和体细胞胚胎发生为濒危物种椭圆叶石蝴蝶（Metabriggsia ovalifolia W. T. Wang）建立了一种高效的双再生途径，该途径是从叶片外植体建立的。当细胞分裂素6-苄基腺嘌呤（BA）或噻二唑素（TDZ）以及生长素吲哚-3-丁酸（IBA）、α-萘乙酸（NAA）和吲哚-3-乙酸（IAA）以2.5μM的浓度应用于基础Murashige和Skoog（MS）培养基时，可以诱导芽的产生。BA和TDZ诱导的不定芽（分别为19.1个/外植体和31.2个/外植体）比NAA（4.6个/外植体）、IBA（5.7个/外植体）或IAA（6.4个/外植体）更多。5-10μM的BA和TDZ可以诱导芽和体细胞胚胎的产生。更高浓度的TDZ（25μM）仅诱导体细胞胚胎（39.8个/外植体）。相同浓度的BA诱导不定芽（23.6个/外植体）和体细胞胚胎（9.7个/外植体）。因此，组织学证据表明，该植物的体细胞胚胎发生需要高浓度的细胞分裂素（BA；TDZ）。体细胞胚胎留在相同培养基上时很容易萌发，但在添加了0.5μM NAA、0.5μM IBA和0.1%活性炭的MS培养基上两周后形成不定根。驯化并转移到托盘中的沙子和蛭石混合物（1:1，v/v）中后，超过93%的植株存活。
