Hussein Gihan M H, Abdel-Rahman Tahany M A, Alwan A H
Gene Transfer Lab, Plant Genetic Transformation Department, Agricultural Genetic Engineering Institute (AGERI), Agricultural Research Center (ARC), Giza, Egypt.
Botany and Microbiolgy Department, Faculty of Science, Cairo University, Giza, Egypt.
3 Biotech. 2016 Dec;6(2):130. doi: 10.1007/s13205-016-0431-9. Epub 2016 Jun 8.
In this study, cell-suspension culture of strawberry (Fragaria × ananassa), cultivars Camarosa, and Sweet Charlie has been established. Embryogenic callus was induced by incubating the in vitro juvenile leaf explants on medium, containing 2-mg/l picloram at dark. Suspension culture was initiated from 4-week-old embryogenic calli in the liquid MS medium with 1-mg/l 2,4-D and 2-mg/l picloram. Suspension culture was maintained by sub-culturing each 3 weeks into a fresh medium. At week 9 after third sub-cultures, torpedo and cotyledonary embryo stages were observed. Embryos were then developed into shoots on medium 1 mg/l of each BA and IBA. Obtained shoots were successfully rooted on 1-mg/ml GA3, 0.5-mg/ml BA, and 1-mg/ml IBA. To enhance the resistance availability in strawberry plants, elicitation was applied by adding the JA and SA elicitors to the suspension culture with two doses (0.5 and 1 mM) individually and in combination, in addition to the fungal homogenate of Macrophomina phasiolena at concentration of 10 spor/ml. The fawrky-1-Camarosa gene, which has defense-related function, was detected in the different elicited strawberry tissues and isolated via RT-PCR. The isolated gene was submitted to GenBank with accession number (KX096885).
在本研究中，已建立了草莓（凤梨草莓）品种卡玛罗莎（Camarosa）和甜查理（Sweet Charlie）的细胞悬浮培养体系。通过将离体幼叶外植体置于含有2毫克/升毒莠定的培养基上，在黑暗条件下诱导胚性愈伤组织。悬浮培养从4周龄的胚性愈伤组织开始，在含有1毫克/升2,4 - D和2毫克/升毒莠定的液体MS培养基中进行。悬浮培养每3周转接至新鲜培养基中进行继代培养以维持。在第三次继代培养后的第9周，观察到鱼雷胚和子叶胚阶段。然后将胚在含有1毫克/升苄氨基腺嘌呤（BA）和1毫克/升吲哚丁酸（IBA）的培养基上发育成芽。获得的芽在含有1毫克/毫升赤霉素（GA3）、0.5毫克/毫升BA和1毫克/毫升IBA的培养基上成功生根。为了提高草莓植株的抗性，除了添加浓度为10个孢子/毫升的菜豆壳球孢真菌匀浆外，还分别以两种剂量（0.5和1毫摩尔）单独及组合添加茉莉酸（JA）和水杨酸（SA）诱导子到悬浮培养物中。在不同诱导处理的草莓组织中检测到具有防御相关功能的fawrky - 1 - Camarosa基因，并通过逆转录聚合酶链反应（RT - PCR）进行分离。分离得到的基因已提交至GenBank，登录号为（KX096885）。
