Eucommia ulmoides, also known as the industrially and medicinally important hardy rubber tree, is the sole species of Eucommiaceae. Nevertheless, its dioecious property hinders sex recognition by traditional morphological observation at very early developmental stages, thus inhibiting breeding and economic cropping. In this study, double-digest restriction site-associated DNA sequencing (ddRAD-seq) was applied to screen sex-linked molecular markers for sex identification and investigation of the sex determination system in 20 male and female E. ulmoides individual plants, respectively. In consequence, five candidate male-specific loci but no female-specific loci were predicated among the 183,752 male and 147,122 female catalogue loci by bioinformatics analysis. Subsequent PCR (polymerase chain reaction) amplification and Sanger sequencing examinations were performed on another 24 individuals, 12 for each sex, from a separate population. One ideal sex-linked locus, MSL4, was identified among the five putative male-specific loci that were found using ddRAD data. MSL4 is 479 bp in length and highly conserved in all the male individuals, suggesting its feature of being stable and repeatable. Our results also indicated that the sex of E. ulmoides is likely determined genetically. In short, this study provides a consistent and reproducible ddRAD marker (MSL4) that is able to discriminate male from female seedlings in E. ulmoides, which will be valuable for rapid breeding practice and better commercial production of this economically important tree.

中文翻译：

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通过ddRAD标记鉴定Hardy橡胶树（Eucommia ulmoides Oliver）中的分子性别。

杜仲（Eucommia ulmoides），也被称为工业上和医学上重要的强硬橡胶树，是杜仲科的唯一品种。然而，其雌雄异株的特性在很早的发育阶段就阻碍了传统形态学观察中的性别识别，从而抑制了育种和经济种植。在这项研究中，双消化限制酶切位点相关的DNA测序（ddRAD-seq）被用于筛选性别相关的分子标记物，以进行性别鉴定和调查20名男性和女性的E. ulmoides中的性别确定系统。分别是单个植物。结果，通过生物信息学分析，确定了183,752个男性和147,122个女性目录基因座中的五个候选男性特定基因座，但没有女性特定基因座。随后对另外24个人进行了PCR（聚合酶链反应）扩增和Sanger测序检查，每个人的性别分别为12。在使用ddRAD数据发现的五个推定的男性特异性基因座中，鉴定出一个理想的性别相关基因座MSL4。MSL4的长度为479 bp，在所有男性个体中高度保守，表明其稳定且可重复的特征。我们的结果还表明E的性别。乌药可能是遗传决定的。简而言之，这项研究提供了一致且可重现的ddRAD标记（MSL4），该标记能够区分E中的雄性和雌性幼苗。ulmoides，对于这种经济上重要的树的快速育种实践和更好的商业生产将是有价值的。