Alligatorweed [Alternanthera philoxeroides (Mart.) Griseb.] is an invasive semiaquatic weed that poses a serious threat to agricultural production and ecological balance worldwide. However, information about genetic factors associated with the adaptation and invasion mechanisms of this species is limited. Screening for appropriate reference genes is important for gene expression and functional analysis research in A. philoxeroides. In this study, 30 candidate genes that showed stable expression in different A. philoxeroides tissues under various treatments in RNA-seq data were chosen to design quantitative real-time PCR (qRT-PCR) primers. After the amplification specificity validation, 25 candidates were selected and further evaluated in a diverse set of A. philoxeroides samples, including leaf, stem, and root tissues under drought, salinity, heat, chilling, five herbicides, and corresponding untreated controls using qRT-PCR. The delta-CT method, geNorm, NormFinder, BestKeeper, and RefFinder algorithms were used to identify stable reference genes from A. philoxeroides samples. Overall, CoA, RFI2, Tubby, SRP19, and V-ATPase were the top five ideal reference genes in all organs and conditions. Tubby and CoA were the most stable reference genes in the leaf/stem; and RFI2, ERprr, and SPR19 were suitable reference genes for the roots. This work provided a foundation for exploring gene expression profiling of A. philoxeroides, especially those adaptation- and invasion-related genes, which may help in management of this invasive weed.

中文翻译：

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鉴定和验证用于短吻鳄草（空心莲子草）定量实时 PCR 研究的参考基因

鳄鱼草 [空心莲子草(Mart.) Griseb.] 是一种侵入性的半水生杂草，对全世界的农业生产和生态平衡构成严重威胁。然而，关于与该物种的适应和入侵机制相关的遗传因素的信息是有限的。筛选合适的参考基因对于基因表达和功能分析研究很重要空心莲子. 在这项研究中，30 个候选基因在不同的基因组中表现出稳定的表达空心莲子选择在 RNA-seq 数据中进行各种处理的组织来设计定量实时 PCR (qRT-PCR) 引物。在扩增特异性验证后，选择了 25 个候选者，并在不同的一组空心莲子样本，包括在干旱、盐分、高温、低温、五种除草剂和相应的未处理对照下使用 qRT-PCR 的叶、茎和根组织。delta-CT 方法、geNorm、NormFinder、BestKeeper 和 RefFinder 算法用于从空心莲子样品。全面的，辅酶A,射频干扰2,胖子,SRP19， 和五-ATP酶是所有器官和条件下的前五个理想参考基因。胖子和辅酶A是叶/茎中最稳定的参考基因；和射频干扰2,ERprr， 和SPR19是适合根的参考基因。这项工作为探索基因表达谱提供了基础一种.空心莲，尤其是那些适应和入侵相关的基因，这可能有助于管理这种入侵杂草。