Castillo Ana M, Sánchez-Díaz Rosa A, Vallés María P
Departamento de Genética y Producción Vegetal, Estación Experimental de Aula Dei, Consejo Superior de Investigaciones Científicas (EEAD-CSIC) Zaragoza, Spain.
Front Plant Sci. 2015 Jun 18;6:402. doi: 10.3389/fpls.2015.00402. eCollection 2015.
Ovary pre-conditioned medium and ovary co-culture increased the efficiency of green doubled haploid plant production in bread wheat anther culture. The positive effect of this medium led to a 6- and 11-fold increase in the numbers of embryos and green plants, respectively, having a greater effect on a medium-low responding cultivar. Ovary genotype and developmental stage significantly affected microspore embryogenesis. By the use of Caramba ovaries it was possible to reach a 2-fold increase in the number of embryos and green plants, and to decrease the rate of albinism. Mature ovaries from flowers containing microspores at a late binucleate stage raised the number of embryos and green plants by 25-46% as compared to immature ovaries (excised from flowers with microspores at a mid-late uninucleate stage). The highest numbers of embryos and green plants were produced when using mature Caramba ovaries. Ovaries from Galeón, Tigre, and Kilopondio cultivars successfully induced microspore embryogenesis at the same rate as Caramba ovaries. Moreover, Tigre ovaries raised the percentage of spontaneous chromosome doubling up to 71%. Attempts were made to identify molecular mechanisms associated to the inductive effect of the ovaries on microspore embryogenesis. The genes TAA1b, FLA26, and WALI6 associated to wheat microspore embryogenesis, the CGL1 gene involved in glycan biosynthesis or degradation, and the FER gene involved in the ovary signaling process were expressed and/or induced at different rates during ovary culture. The expression pattern of FLA26 and FER could be related to the differences between genotypes and developmental stages in the inductive effect of the ovary. Our results open opportunities for new approaches to increase bread wheat doubled haploid production by anther culture, and to identify the functional components of the ovary inductive effect on microspore embryogenesis.
卵巢预处理培养基和卵巢共培养提高了面包小麦花药培养中绿色双单倍体植株的生产效率。这种培养基的积极作用分别使胚和绿色植株的数量增加了6倍和11倍，对中低反应型品种的影响更大。卵巢基因型和发育阶段显著影响小孢子胚胎发生。使用卡兰巴（Caramba）品种的卵巢，可使胚和绿色植株的数量增加2倍，并降低白化率。与未成熟卵巢（从单核中晚期小孢子的花中切取）相比，含有双核晚期小孢子的花中的成熟卵巢使胚和绿色植株的数量增加了25%-46%。使用成熟的卡兰巴卵巢时，产生的胚和绿色植株数量最多。来自加莱昂（Galeón）、蒂格雷（Tigre）和基洛庞德奥（Kilopondio）品种的卵巢成功诱导小孢子胚胎发生的速率与卡兰巴卵巢相同。此外，蒂格雷卵巢使自发染色体加倍的百分比提高到了71%。人们试图确定与卵巢对小孢子胚胎发生的诱导作用相关的分子机制。与小麦小孢子胚胎发生相关的TAA1b、FLA26和WALI6基因，参与聚糖生物合成或降解的CGL1基因，以及参与卵巢信号传导过程的FER基因，在卵巢培养过程中以不同速率表达和/或被诱导。FLA26和FER的表达模式可能与卵巢诱导作用在基因型和发育阶段上的差异有关。我们的研究结果为通过花药培养提高面包小麦双单倍体产量的新方法以及确定卵巢对小孢子胚胎发生诱导作用的功能成分提供了机会。
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