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.

The storage root of alligatorweed [Alternanthera philoxeroides(Mart.) Griseb.] growing in terrestrial habitats is an important metamorphic organ for its propagation, overwintering, and spread. However, the regulatory mechanism adventitious root expansion to form storage roots is still unclear. To reveal the changes accompanying the root-swelling process, we quantified sugar, soluble protein, and phytohormone content in adventitious and storage roots. Results demonstrated that sucrose, fructose, and soluble protein increased in storage roots, whereas abscisic acid (ABA), indoleacetic acid (IAA), brassinosteroid (BR), gibberellin, jasmonic acid, and cytokinin (trans-zeatin [tZ] and isopentenyladenine [iP] and the corresponding ribosidestZR and iPR).tZ-type (tZR andtZ) content decreased, suggesting the involvement of sugars and hormones in the formation of storage roots. To further reveal the molecular basis ofA.philoxeroides’s ability to form storage roots and provide candidate genes for molecular function analyses, we assembled a de novo transcriptome ofA.philoxeroidesbased on four sets of RNA-sequencing data. According to functional annotation and expression profiling, 42 unigenes involved in sucrose synthesis and hydrolysis were identified, in addition to 70, 58, and 78 unigenes in ABA, BR, and IAA signal transduction, respectively. The quantitative reverse transcriptase polymerase chain reaction analysis revealed 21 unigenes involved in sugar metabolism and hormone signal transduction were differentially expressed during the formation of storage roots. These results revealed metabolic changes during the formation of storage roots and provide candidate genes involved in sugar and phytohormone metabolism inA. philoxeroides.