Tissue culture tools for selenium hyperaccumulator Neptunia amplexicaulis for development in phytoextraction.

Neptunia amplexicaulis is an herbaceous legume endemic to the Richmond area in central Queensland, Australia and is one of the strongest known Selenium hyperaccumulators on earth, showing significant potential to be utilised in Se phytoextraction applications. Here a protocol was established for in vitro micropropagation of Se hyperaccumulator N. amplexicaulis using nodal segments from in vitro-germinated seedlings. Shoot multiplication was achieved on Murashige and Skoog (MS) basal media supplemented with various concentrations of 6-Benzylaminopurine (BA) (1.0, 2.0, 3.0 mg L-1) alone or in combination with low levels of Naphthaleneacetic acid (NAA) (0.1, 0.2, 0.3 mg L-1), with 2.0 mg L-1 BA + 0.2 mg L-1 NAA found to be most effective. Elongated shoots were rooted in vitro using NAA, with highest root induction rate of 30% observed at 0.2 mg L-1 NAA. About 95% of the in vitro rooted shoots survived acclimatization. Clonally propagated plantlets were dosed with selenate/selenite solution and assessed for Se tissue concentrations using Inductively Coupled Plasma Atomic Emission Spectroscopy (ICP-AES) and found to retain their ability to hyperaccumulate. The protocol developed for this study has potential to be optimised for generating clonal plants of N. amplexicaulis for use in research and phytoextraction industry applications.

Chromosome-level genome map provides insights into diverse defense mechanisms in the medicinal fungus Ganoderma sinense.

Fungi have evolved powerful genomic and chemical defense systems to protect themselves against genetic destabilization and other organisms. However, the precise molecular basis involved in fungal defense remain largely unknown in Basidiomycetes. Here the complete genome sequence, as well as DNA methylation patterns and small RNA transcriptomes, was analyzed to provide a holistic overview of secondary metabolism and defense processes in the model medicinal fungus, Ganoderma sinense. We reported the 48.96 Mb genome sequence of G. sinense, consisting of 12 chromosomes and encoding 15,688 genes. More than thirty gene clusters involved in the biosynthesis of secondary metabolites, as well as a large array of genes responsible for their transport and regulation were highlighted. In addition, components of genome defense mechanisms, namely repeat-induced point mutation (RIP), DNA methylation and small RNA-mediated gene silencing, were revealed in G. sinense. Systematic bioinformatic investigation of the genome and methylome suggested that RIP and DNA methylation combinatorially maintain G. sinense genome stability by inactivating invasive genetic material and transposable elements. The elucidation of the G. sinense genome and epigenome provides an unparalleled opportunity to advance our understanding of secondary metabolism and fungal defense mechanisms.

Enhanced seed oil content by overexpressing genes related to triacylglyceride synthesis.

Oilseed rape (Brassica napus) is one of the most important oilseed crops globally. To meet increasing demand for oil-based products, the ability to enhance desirable oil content in the seed is required. This study assessed the capability of five genes in the triacylglyceride (TAG) synthesis pathway to enhance oil content. The genes BnGPDH, BnGPAT, BnDGAT, ScGPDH and ScLPAAT were overexpressed separately in a tobacco (Nicotiana benthamiana) model system, and simultaneously by pyramiding in B. napus, under the control of a seed specific Napin promoter. ScLPAAT transgenic plants showed a significant increase of 6.84% to 8.55% in oil content in tobacco seeds, while a ~4% increase was noted for BnGPDH and BnGPAT transgenic seeds. Seed-specific overexpression of all four genes in B. napus resulted in as high a 12.57% to 14.46% increased in seed oil content when compared to WT, equaling close to the sum of the single-gene overexpression increases in tobacco. Taken together, our study demonstrates that BnGPDH, BnGPAT and ScLPAAT may effectively increase seed oil content, and that simultaneous overexpression of these in transgenic B. napus may further enhance the desirable oil content relative to single-gene overexpressors.

Genome-wide identification of phenolic acid biosynthetic genes in Salvia miltiorrhiza.

MAIN CONCLUSION: Twenty-nine genes related to phenolic acid biosynthesis were identified in the Salvia miltiorrhiza genome. Nineteen of these are described for the first time, with ten genes experimentally correlating to phenolic acid biosynthesis. Vast stores of secondary metabolites exist in plants, many of which possess biological activities related to human health. Phenolic acid derivatives are a class of valuable bioactive pharmaceuticals abundant in the widely used Chinese medicinal herb, Salvia miltiorrhiza. The biosynthetic pathway for phenolic acids differs in this species from that of other investigated plants. However, the molecular basis for this is unknown, with systematic analysis of the genes involved not yet performed. As the first step towards unraveling this complex biosynthetic pathway in S. miltiorrhiza, the current genome assembly was searched for putatively involved genes. Twenty-nine genes were revealed, 19 of which are described here for the first time. These include 15 genes predicted in the phenylpropanoid pathway; seven genes in the tyrosine-derived pathway; six genes encoding putative hydroxycinnamoyltransferases, and one CYP98A, namely CYP98A78. The promoter regions, gene structures and expression patterns of these genes were examined. Furthermore, conserved domains and phylogenetic relationships with homologous proteins in other species were revealed. Most of the key enzymes, including 4-coumarate: CoA ligase, 4-hydroxyphenylpyruvate reductase and hydroxycinnamoyltransferase, were found in multiple copies, each exhibiting different characteristics. Ten genes putatively involved in rosmarinic acid biosynthesis are also described. These findings provide a foundation for further analysis of this complex and diverse pathway, with potential to enhance the synthesis of water-soluble medicinal compounds in S. miltiorrhiza.