Cymbidium has been artificially domesticated for centuries in Asia, which produced numerous cultivated varieties. Flowers with stamenoid tepals or those with multiple tepals have been found in different species of Cymbidium; however, the molecular basis controlling the formation of these phenotypes is still largely unknown. Previous work demonstrated that AGAMOUS/AG lineage MADS genes function in floral meristem determinacy as well as in reproductive organs development in both dicots and monocots, indicating a possible relationship with the origin of two flower varieties in Cymbidium. Here, we characterized and analyzed two AG lineage paralogues, CsAG1 and CsAG2, from Cymbidium sinense, both of which were highly expressed in the gynostemium column of a standard C. sinense. Interestingly, we detected ectopic expression of CsAG1 rather than CsAG2 in all floral organs of a stamenoid-tepal variety and significant down-regulation of CsAG1 in a variety with multiple tepals. Over-expression of CsAG1 in wild type Arabidopsis resulted in petal-to-stamen homeotic conversion, suggesting a conserved C-function of CsAG1 in the development of Cymbidium flower. Altogether, our results supported a hypothesis that disruption of a single AG-like factor would be associated with the formation of two domesticated varieties in C. sinense.

Phosphoenolpyruvate carboxylase (PEPC) is a ubiquitous cytosolic enzyme that catalyzes the irreversible β-carboxylation of phosphoenolpyruvate (PEP) in presence of HCO₃⁻ to produce oxaloacetate (OAA) during carbon fixation and photosynthesis. It is well accepted that PEPC genes are expressed in plants upon stress. PEPC also supports the biosynthesis of biocompatible osmolytes in many plant species under osmotic stress. There are five isoforms of PEPC found in peanut (Arachis hypogaea L.), namely, AhPEPC1, AhPEPC2, AhPEPC3, AhPEPC4, and AhPEPC5. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the gene expression patterns of these AhPEPC genes were different in mature seeds, stems, roots, flowers, and leaves. The expression of all the plant type PEPC (PTPCs) (AhPEPC1, AhPEPC2, AhPEPC3, and AhPEPC4) was relatively high in roots, while the bacterial type PEPC (BTPC) (AhPEPC5) showed a remarkable expression level in flowers. Principal component analysis (PCA) result showed that AhPEPC3 and AhPEPC4 are correlated with each other, indicating comparatively associations with roots, and AhPEPC5 have a very close relationship with flowers. In order to investigate the function of these AhPEPCs, the fragments of these five AhPEPC cDNA were cloned and expressed in Escherichia coli (E. coli). The recombinant proteins contained a conserved domain with a histidine site, which is important for enzyme catalysis. Results showed that protein fragments of AhPEPC1, AhPEPC2, and AhPEPC5 had remarkable expression levels in E. coli. These three recombinant strains were more sensitive at pH 9.0, and recombinant strains carrying AhPEPC2 and AhPEPC5 fragments exhibited more growth than the control strain with the presence of PEG6000. Our findings showed that the expression of the AhPEPC fragments may enhance the resistance of transformed E. coli to osmotic stress.

Orchid flowers have evolved in concert with pollinators to form highly specialized structures resulting in zygomorphy. In dicotyledons, it is widely accepted that CYC-like genes are involved in the dorsoventral polarity establishment of flowers, which determines the development of zygomorphic flowers. However, the function of TCP transcription factors involved in orchid floral development is rarely known. Here, we found 15 unigenes with TCP domain (EpTCPs) from the previously reported Erycina pusilla unigene database. The expression patterns of EpTCPs in various tissues and different floral organs were successively detected by quantitative real-time PCR. The results revealed that the CYC-like gene (EpTCP25) and CIN-like genes (EpTCP11 and EpTCP26) were highly expressed in inflorescences but lowly expressed in leaves and roots. What is more, these three genes were expressed relatively high in the dorsal labellum, and EpTCP26 showed differential expression along the dorsoventral polarity of tepals, which was high in the dorsal and low in the ventral. Ectopic expression of EpTCP25 in Arabidopsis repressed primary root growth and delayed flowering. EpTCP26 overexpression in Arabidopsis promoted primary root growth and leaf growth. In contrast, EpTCP11 overexpression repressed primary root growth and changed the radially symmetric flower to a bilaterally symmetric flower by inhibiting the elongation of one or two adjacent petals. In addition, the homeotic transition of floral organs is generated when these genes are ectopically expressed in Arabidopsis, suggesting their roles in floral morphogenesis. Altogether, our results indicate that CIN-like genes would be associated with the unique flower pattern development of Erycina pusilla.

Mikania cordata, the only native congener of the invasive weed Mikania micrantha in China, is an ideal species for comparative study to reveal the invasion mechanism. However, its genome resources are lagging far behind its congener, which limits the comparative genomic analysis. Our goal is to characterize the genome of M. cordata by next-generation sequencing and propose a scheme for long-read genome sequencing. Previous studies have shown that the genomic resources of the host plant would be affected by the endophytic microbial DNA. An aseptic sample of M. cordata will ensure the proper genome in downstream analysis. Because endophytes are ubiquitous in the greenhouse-grown M. cordata, the in vitro culture with cefotaxime or timentin treatment was undertaken to obtain the aseptic plantlets. The in vivo mother plant and in vitro plantlets were used to survey the genome. The microbial contamination in M. cordata was recognized by blast search and eliminated from the raw reads. The decontaminated sequencing reads were used to predict the genome size, heterozygosity, and repetitive rate. The in vivo plant was so contaminated that microbes occupied substantial sequencing resources and misled the scaffold assembly. Compared with cefotaxime, treatment with timentin performed better in cultivating robust in vitro plantlets. The survey result from the in vitro plantlets was more accurate due to low levels of contamination. The genome size was estimated to be 1.80 Gb with 0.50% heterozygosity and 78.35% repetitive rate. Additionally, 289,831 SSRs were identified in the genome. The genome is heavily contaminated and repetitive; therefore, the in vitro culture technique and long-read sequencing technology are recommended to generate a high-quality and highly contiguous genome.