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Metabolic engineering of heterologous hosts requires coexpression of multiple genes, often more than ten for a single pathway. Traditional approaches to create genetic constructs for this purpose are highly inflexible and suffer from very low throughput. In this book chapter, we describe a powerful method to overcome this bottleneck, namely, combinatorial co-expression in the Australian tobacco plant Nicotiana benthamiana. This system is based on Agrobacterium tumefaciens-mediated transient gene expression, often called agroinfiltration. Herein, instead of creating complex multigenic constructs, coexpression is achieved by simply mixing different plasmid-bearing Agrobacterium strains without the need for different selection markers. We present a practical guide for coexpressing multiple biosynthetic genes followed by GC-MS analysis, using production of the plant triterpene β-amyrin as an example. Our chapter provides a guideline to harness the potential of this versatile expression system in the natural product community for studying and engineering metabolic pathways., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Fusarium goolgardi, isolated from the grass tree Xanthorrhoea glauca in natural ecosystems of Australia, is closely related to fusaria that produce a subgroup of trichothecene (type A) mycotoxins that lack a carbonyl group at carbon atom 8 (C-8). Mass spectrometric analysis revealed that F. goolgardi isolates produce type A trichothecenes, but exhibited one of two chemotypes. Some isolates (50%) produced multiple type A trichothecenes, including 4,15-diacetoxyscirpenol (DAS), neosolaniol (NEO), 8-acetylneosolaniol (Ac-NEO) and T-2 toxin (DAS-NEO-T2 chemotype). Other isolates (50%) produced only DAS (DAS chemotype). In the phylogenies inferred from DNA sequences of genes encoding the RNA polymerase II largest (RPB1) and second largest (RPB2) subunits as well as the trichothecene biosynthetic genes (TRI), F. goolgardi isolates were resolved as a monophyletic clade, distinct from other type A trichothecene-producing species. However, the relationships of F. goolgardi to the other species varied depending on whether phylogenies were inferred from RPB1 and RPB2, the 12-gene TRI cluster, the two-gene TRI1-TRI16 locus, or the single-gene TRI101 locus. Phylogenies based on different TRI loci resolved isolates with different chemotypes into distinct clades, even though only the TRI1-TRI16 locus is responsible for structural variation at C-8. Sequence analysis indicated that TRI1 and TRI16 are functional in F. goolgardi isolates with the DAS-NEO-T2 chemotype, but non-functional in isolates with DAS chemotype due to the presence of premature stop codons caused by a point mutation.

The cyanobacterium Dolichospermum circinale (formerly Anabaena circinalis ) is responsible for neurotoxic saxitoxin-producing blooms in Australia. Previous studies have reported distinct isolates of toxic D. circinale producing different saxitoxin analogues at varying amounts, but the mechanisms responsible remain poorly understood. To assess the characteristics that may be responsible for this variance, a morphological, molecular and chemical survey of 28 Anabaena isolates was conducted. Morphological characteristics, presence or absence of saxitoxin biosynthetic genes and toxin amount and profile were assessed. The 28 isolates were collected from 16 locations. A correlation between the size of the isolates and its reported toxicity or geographical location could not be found. Molecular screening for the presence of several sxt genes revealed eight out of the 28 strains harboured the sxt gene cluster and all tailoring genes except sxtX . Furthermore, the presence of PSTs was correlated with the presence of the sxt cluster using quantitative pre-column oxidation high performance liquid chromatography with fluorescence detection (HPLC-FLD) and LC-MS/MS. Interestingly, isolates differed in the amount and type of toxins produced, with the eight toxic strains containing the core and tailoring biosynthetic genes while non-toxic strains were devoid of these genes. Moreover, the presence of sxt tailoring genes in toxic strains correlated with the biosynthesis of analogues. A greater understanding of toxin profile/quantity from distinct sites around Australia will aid the management of these at-risk areas and provide information on the molecular control or physiological characteristics responsible for toxin production. [ABSTRACT FROM AUTHOR]

Wollemi pine (Wollemia nobilis (Jones, Hill, Allen)) is a living fossil, known only through fossil records until its 1994 discovery in Australia. Wollemi Pine is closely related to Norfolk Island pine (Araucaria heterophylla (Salisb.) Franco), making it an interesting system to study metabolic evolution. We employed untargeted liquid chromatography mass spectrometry-based (LC-MS) metabolomics, with chemometrics, pathway analysis, and our novel plant growth regulator (PGR) putative identification tool (HormonomicsDB), to explore the metabolomes of both species. We identified PGR conjugates and found cytokinin, zeatin, and brassinosteroid pathways to be overrepresented in Wollemi pine tissues, suggesting these PGRs play an important role in its survival. Melatonin, a proposed ancient and stress-related PGR, was not identified in untargeted analysis of either species. Based on our untargeted results, we employed targeted LC-MS to quantify brassinolide and confirm the absence or presence of melatonin in Wollemi and Norfolk Island pine. Both species had similar concentrations of brassinolide, while Wollemi pine had significantly higher melatonin levels than Norfolk Island pine. High levels of melatonin and brassinolide in Wollemi pine supports the hypothesis that these are ancient PGRs that conferred an evolutionary advantage in Wollemi, allowing it to persist. [ABSTRACT FROM AUTHOR]

Key Message: The vacuolar processing enzyme gene TaVPE3cB is identified as a candidate gene for a QTL of wheat pith-thickness on chromosome 3B by BSR-seq and differential expression analyses. The high pith-thickness (PT) of the wheat stem could greatly enhance stem mechanical strength, especially the basal internodes which support the heavier upper part, such as upper stems, leaves and spikes. A QTL for PT in wheat was previously discovered on 3BL in a double haploid population of 'Westonia' × 'Kauz'. Here, a bulked segregant RNA-seq analysis was applied to identify candidate genes and develop associated SNP markers for PT. In this study, we aimed at screening differentially expressed genes (DEGs) and SNPs in the 3BL QTL interval. Sixteen DEGs were obtained based on BSR-seq and differential expression analyses. Twenty-four high-probability SNPs in eight genes were identified by comparing the allelic polymorphism in mRNA sequences between the high PT and low PT samples. Among them, six genes were confirmed to be associated with PT by qRT-PCR and sequencing. A putative vacuolar processing enzyme gene TaVPE3cB was screened out as a potential PT candidate gene in Australian wheat 'Westonia'. A robust SNP marker associated with TaVPE3cB was developed, which can assist in the introgression of TaVPE3cB.b in wheat breeding programs. In addition, we also discussed the function of other DEGs which may be related to pith development and programmed cell death (PCD). A five-level hierarchical regulation mechanism of stem pith PCD in wheat was proposed., (© 2023. The Author(s).)

Many plants of Dendrobium genus are precious traditional herbs with high commercial value and excellent medicinal effects. They are perennial aerophytes or epiphytes of terrestrial orchids growing on cliffs and tree trunks covered with mosses in forests throughout the tropical and subtropical Asia and eastern Australia. The stem contains a variety of bioactive components, including polysaccharides and alkaloids, with strong antioxidant, neuroprotective, and immunomodulatory effects. Great attention has been drawn to the Dendrobium genus regarding its medicinal effectiveness, and the related researches have been accumulating rapidly in recent years. The bioactive components are mainly the intermediates or final products produced in specialized metabolite biosynthesis. Thus far, the activity, molecular structure, and composition of major medicinal ingredients have been partially elucidated, and the sequencing of several transcriptomes has been starting to shed new light on the biosynthesis regulation mechanism. This paper reviewed the advances of researches concerning the biosynthetic pathways of medicinal specialized metabolites from Dendrobium , especially the large number of related genes, with the hope of further promoting the development and utilization of those components and correspondingly protecting the Dendrobium resources in more effective ways. [ABSTRACT FROM AUTHOR]

Background: Eremophila R.Br. (Scrophulariaceae) is a diverse genus of plants with species distributed across semi-arid and arid Australia. It is an ecologically important genus that also holds cultural significance for many Indigenous Australians who traditionally use several species as sources of medicines. Structurally unusual diterpenoids, particularly serrulatane and viscidane-types, feature prominently in the chemical profile of many species and recent studies indicate that these compounds are responsible for much of the reported bioactivity. We have investigated the biosynthesis of diterpenoids in three species: Eremophila lucida, Eremophila drummondii and Eremophila denticulata subsp. trisulcata. Results: In all studied species diterpenoids were localised to the leaf surface and associated with the occurrence of glandular trichomes. Trichome-enriched transcriptome databases were generated and mined for candidate terpene synthases (TPS). Four TPSs with diterpene biosynthesis activity were identified: ElTPS31 and ElTPS3 from E. lucida were found to produce (3Z,7Z,11Z)-cembratrien-15-ol and 5-hydroxyviscidane, respectively, and EdTPS22 and EdtTPS4, from E. drummondii and E. denticulata subsp. trisulcata, respectively, were found to produce 8,9-dihydroserrulat-14-ene which readily aromatized to serrulat-14-ene. In all cases, the identified TPSs used the cisoid substrate, nerylneryl diphosphate (NNPP), to form the observed products. Subsequently, cis-prenyl transferases (CPTs) capable of making NNPP were identified in each species. Conclusions: We have elucidated two biosynthetic steps towards three of the major diterpene backbones found in this genus. Serrulatane and viscidane-type diterpenoids are promising candidates for new drug leads. The identification of an enzymatic route to their synthesis opens up the possibility of biotechnological production, making accessible a ready source of scaffolds for further modification and bioactivity testing. [ABSTRACT FROM AUTHOR]

Armillaria root rot is a fungal disease that affects a wide range of trees and crops around the world. Despite being a widespread disease, little is known about the plant molecular responses towards the pathogenic fungi at the early phase of their interaction. With recent research highlighting the vital roles of metabolites in plant root–microbe interactions, we sought to explore the presymbiotic metabolite responses of Eucalyptus grandis seedlings towards Armillaria luteobuablina, a necrotrophic pathogen native to Australia. Using a metabolite profiling approach, we have identified threitol as one of the key metabolite responses in E. grandis root tips specific to A. luteobubalina that were not induced by three other species of soil‐borne microbes of different lifestyle strategies (a mutualist, a commensalist, and a hemi‐biotrophic pathogen). Using isotope labelling, threitol detected in the Armillaria‐treated root tips was found to be largely derived from the fungal pathogen. Exogenous application of d‐threitol promoted microbial colonization of E. grandis and triggered hormonal responses in root cells. Together, our results support a role of threitol as an important metabolite signal during eucalypt‐Armillaria interaction prior to infection thus advancing our mechanistic understanding on the earliest stage of Armillaria disease development. Comparative metabolomics of eucalypt roots interacting with a range of fungal lifestyles identified threitol enrichment as a specific characteristic of Armillaria pathogenesis. Our findings suggest that threitol acts as one of the earliest fungal signals promoting Armillaria colonization of roots. Comparative metabolomics of eucalypt roots interacting with different fungi identified threitol enrichment as a specific characteristic of Armillaria pathogenesis. Our findings suggest that threitol acts as one of the earliest fungal signals promoting Armillaria colonization of roots. [ABSTRACT FROM AUTHOR]

Main conclusion: The HEUKCHEEM gene plays an important role in spine color formation. A white spine occurs due to two mutations in HEUKCHEEM and is closely related to the regional distribution of these mutants. Mapping analysis revealed that the HEUKCHEEM gene is co-segregated with the B locus in the regulation of black spine color development in cucumber fruit. HEUKCHEEM induced the expression of the genes involved in the anthocyanin biosynthetic pathway, leading to the accumulation of anthocyanins in black spines. The transiently over-expressed HEUKCHEEM in cucumber and tobacco plants enhanced the expression of anthocyanin biosynthesis-related genes, leading to anthocyanin accumulation. However, two mutations—insertion of the 6994 bp mutator-like transposable element (MULE) sequence into the second intron and one single-nucleotide polymorphism (SNP) of C to T in the second exon of HEUKCHEEM—were identified in white spines, leading to no accumulation of anthocyanin biosynthesis-related gene transcripts and anthocyanins. Furthermore, association analysis using 104 cucumber accessions with different geographical origins revealed that the types of mutations in HEUKCHEEM are strongly linked to geographical origins. The MULE insertion is found extensively in cucumbers with white spines in East Asia and Australia. However, cucumbers with white spines in other areas could be significantly influenced by a single SNP mutation. Our results provide fundamental information on spine color development in cucumber fruits and spine color-based cucumber breeding programs. [ABSTRACT FROM AUTHOR]

Colletotrichum tanaceti is an emerging foliar fungal pathogen of commercially grown pyrethrum (Tanacetum cinerariifolium). Despite being reported consistently from field surveys in Australia, the molecular basis of pathogenicity of C. tanaceti on pyrethrum is unknown. Herein, the genome of C. tanaceti (isolate BRIP57314) was assembled de novo and annotated using transcriptomic evidence. The inferred putative pathogenicity gene suite of C. tanaceti comprised a large array of genes encoding secreted effectors, proteases, CAZymes and secondary metabolites. Comparative analysis of its putative pathogenicity gene profiles with those of closely related species suggested that C. tanaceti likely has additional hosts to pyrethrum. The genome of C. tanaceti had a high repeat content and repetitive elements were located significantly closer to genes inferred to influence pathogenicity than other genes. These repeats are likely to have accelerated mutational and transposition rates in the genome, resulting in a rapid evolution of certain CAZyme families in this species. The C. tanaceti genome showed strong signals of Repeat Induced Point (RIP) mutation which likely caused its bipartite nature consisting of distinct gene-sparse, repeat and A-T rich regions. Pathogenicity genes within these RIP affected regions were likely to have a higher evolutionary rate than the rest of the genome. This “two-speed” genome phenomenon in certain Colletotrichum spp. was hypothesized to have caused the clustering of species based on the pathogenicity genes, to deviate from taxonomic relationships. The large repertoire of pathogenicity factors that potentially evolve rapidly due to the plasticity of the genome, indicated that C. tanaceti has a high evolutionary potential. Therefore, C. tanaceti poses a high-risk to the pyrethrum industry. Knowledge of the evolution and diversity of the putative pathogenicity genes will facilitate future research in disease management of C. tanaceti and other Colletotrichum spp. [ABSTRACT FROM AUTHOR]

When wetted, Plantago seeds become covered with a polysaccharide-rich gel called mucilage that has value as a food additive and bulking dietary fibre. Industrially, the dry husk layer that becomes mucilage, called psyllium, is milled off Plantago ovata seeds, the only commercial-relevant Plantago species, while the residual inner seed tissues are either used for low value animal feed or discarded. We suggest that this practice is potentially wasting a highly nutritious resource and here describe the use of histological, physicochemical, and chromatographic analyses to compare whole seed composition/characteristics of P. ovata with 11 relatives already adapted to harsh Australian conditions that may represent novel commercial crop options. We show that substantial interspecific differences in mucilage yield and macromolecular properties are mainly a consequence of differences in heteroxylan and pectin composition and probably represent wide differences in hydrocolloid functionality that can be exploited in industry. We also show that non-mucilage producing inner seed tissues contain a substantial mannan-rich endosperm, high in fermentable sugars, protein, and fats. Whole seed Plantago flour, particularly from some species obtained from harsh Australian environments, may provide improved economic and health benefits compared to purified P. ovata psyllium husk, by retaining the functionality of the seed mucilage and providing additional essential nutrients.

Rationale, aims and objectives Declining capacity to function safely and independently in the community is a manifestation of ageing. Multiple measures are used to define and predict functional decline. This paper explores the use of partial least squares ( PLS) analysis to understand the interaction between overtime measures of functional decline. Methods Comprehensive information was captured on individuals aged 65+ who presented at a large metropolitan Australian hospital Emergency Department ( ED) for a complaint, which did not result in a hospital admission. They were followed-up by telephone 1 and 3 months, post- ED discharge. Information was collected each time on home situations (living alone, using community services, using a gait aid and having a formal carer), recent falls, hospitalizations and instrumental activities of daily living ( IADLs). PLS analysis was applied to identify overtime relationships between measures. Results Valid information was provided by 147 individuals at all three time points. The eight individual IADL items clustered clearly around physical, mental or combined mental and physical IADLs. These clusters were strongly related to increased use of community supports, gait aid and carer, and living alone. The relationship was less convincingly for hospitalizations, and falls were not explained well. Conclusion PLS analysis offers a novel and comprehensive way of analysing complex health data, which allows sense to be made of relationships over a 3-month period. This analysis provides a better understanding of declining function over time, than could be provided by current health modelling methods. [ABSTRACT FROM AUTHOR]

The Cylindrospermopsis raciborskii population from Brazilian freshwater is known to produce saxitoxin derivatives (STX), while cylindrospermopsin (CYN), which is commonly detected in isolates from Australia and Asia continents, has thus far not been detected in South American strains. However, during the investigation for the presence of cyrA, cyrB, cyrC and cyrJ CYN synthetase genes in the genomes of four laboratory-cultured C. raciborskii Brazilian strains, the almost complete cyrA gene sequences were obtained for all strains, while cyrB and cyrC gene fragments were observed in two strains. These nucleotide sequences were translated into amino acids, and the predicted protein functions and domains confirmed their identity as CYN synthetase genes. Attempts to PCR amplify cyrJ gene fragments from the four strains were unsuccessful. Phylogenetic analysis grouped the nucleotide sequences together with their homologues found in known CYN synthetase clusters of C. raciborskii strains with high bootstrap support. In addition, fragments of sxtA, sxtB and sxtI genes involved in STX production were also obtained. Extensive LC-MS analyses were unable to detect CYN in the cultured strains, whereas the production of STX and its analogues was confirmed in CENA302, CENA305 and T3. To our knowledge, this is the first study reporting the presence of cyr genes in South American strains of C. raciborskii and the presence of sxt and cyr genes in a single C. raciborskii strain. This discovery suggests a shift in the type of cyanotoxin production over time of South American strains of C. raciborskii and contributes to the reconstruction of the evolutionary history and diversification of cyanobacterial toxins. [ABSTRACT FROM AUTHOR]

Lipopolysaccharide (LPS) is one of the most important virulence and antigenic components of Burkholderia pseudomallei, the causative agent of melioidosis. LPS diversity in B. pseudomallei has been described as typical, atypical or rough, based upon banding patterns on SDS-PAGE. Here, we studied the genetic and molecular basis of these phenotypic differences. Bioinformatics was used to determine the diversity of genes known or predicted to be involved in biosynthesis of the O-antigenic moiety of LPS in B. pseudomallei and its near-relative species. Multiplex-PCR assays were developed to target diversity of the O-antigen biosynthesis gene patterns or LPS genotypes in B. pseudomallei populations. We found that the typical LPS genotype (LPS genotype A) was highly prevalent in strains from Thailand and other countries in Southeast Asia, whereas the atypical LPS genotype (LPS genotype B) was most often detected in Australian strains (∼13.8%). In addition, we report a novel LPS ladder pattern, a derivative of the atypical LPS phenotype, associated with an uncommon O-antigen biosynthesis gene cluster that is found in only a small B. pseudomallei sub-population. This new LPS group was designated as genotype B2. We also report natural mutations in the O-antigen biosynthesis genes that potentially cause the rough LPS phenotype. We postulate that the diversity of LPS may correlate with differential immunopathogenicity and virulence among B. pseudomallei strains. Author Summary: Burkholderia pseudomallei is an environmental Gram-negative bacterium and the cause of melioidosis, an often life-threatening disease affecting people in Southeast Asia and northern Australia. Melioidosis is usually contracted by bacterial inoculation, ingestion or inhalation. Effective vaccines for melioidosis are currently unavailable. This organism contains a large genome, which varies greatly among strains due to a high frequency of genetic recombination. We report here on diversity of lipopolysaccharides (LPS) in this species, a major component of the bacterial outer membrane and a known immunogenic virulence factor. We developed LPS genotyping techniques to study frequency of two major LPS types, known as typical and atypical LPS, in B. pseudomallei strains collected from two endemic regions: Southeast Asia and Northern Australia. LPS genotype variation differed among B. pseudomallei populations. During the investigation, we discovered a new LPS genotype in a sub-population group of B. pseudomallei in Australia. We postulate that such differences are likely to be associated with variable immunopathogenicity and clinical presentation in the human host. [ABSTRACT FROM AUTHOR]

Melioidosis, a febrile illness with disease states ranging from acute pneumonia or septicaemia to chronic abscesses, was first documented by Whitmore & Krishnaswami (1912) . The causative agent, Burkholderia pseudomallei, was subsequently identified as a motile, gram-negative bacillus, which is principally an environmental saprophyte. Melioidosis has become an increasingly important disease in endemic areas such as northern Thailand and Australia ( Currie et al., 2000 ). This health burden, plus the classification of B. pseudomallei as a category B biological agent ( Rotz et al., 2002 ), has resulted in an escalation of research interest. This review focuses on the molecular and cellular basis of pathogenesis in melioidosis, with a comprehensive overview of the current knowledge on how B. pseudomallei can cause disease. The process of B. pseudomallei movement from the environmental reservoir to attachment and invasion of epithelial and macrophage cells and the subsequent intracellular survival and spread is outlined. Furthermore, the diverse assortment of virulence factors that allow B. pseudomallei to become an effective opportunistic pathogen, as well as to avoid or subvert the host immune response, is discussed. With the recent increase in genomic and molecular studies, the current understanding of the infection process of melioidosis has increased substantially, yet, much still remains to be elucidated. [ABSTRACT FROM AUTHOR]

Domestication has resulted in a loss of genetic diversity in our major food crops, leading to susceptibility to biotic and abiotic stresses linked with climate change. Crop wild relatives (CWR) may provide a source of novel genes potentially important for re-gaining climate resilience. Sorghum bicolor is an important cereal crop with wild relatives that are endemic to Australia. Sorghum bicolor is cyanogenic, but the cyanogenic status of wild Sorghum species is not well known. In this study, leaves of wild species endemic in Australia are screened for the presence of the cyanogenic glucoside dhurrin. The direct measurement of dhurrin content and the potential for dhurrin-derived HCN release (HCNp) showed that all the tested Australian wild species were essentially phenotypically acyanogenic. The unexpected low dhurrin content may reflect the variable and generally nutrient-poor environments in which they are growing in nature. Genome sequencing of six CWR and PCR amplification of the CYP79A1 gene from additional species showed that a high conservation of key amino acids is required for correct protein function and dhurrin synthesis, pointing to the transcriptional regulation of the cyanogenic phenotype in wild sorghum as previously shown in elite sorghum. [ABSTRACT FROM AUTHOR]

Brassica napus is cultivated largely as a winter or semi‐winter form in Europe and Asia, respectively, whereas spring‐sown rape types are more adapted to the climatic conditions in Canada, northern Europe, and Australia. This plant, the third most important oil crop in the world after maize and oil palm, has had a unique history among those crops that have been targets of breeding and genetic modification. Although agronomy and genetic manipulations of B. napus plants to create seed with substantially no erucic acid content in the seed oil and meal largely free from aliphatic glucosinolates were major breakthroughs, opportunities for further advances in the reduction of antinutritive compounds/enhancement of bioactive compounds, in oil and meal health safety and quality, as well as in agronomic performance and pest resistance, are still greater today than they have ever been. Understanding and application of genetic control for key agronomic traits in B. napus, such as seed oil composition and concentration as well as their environmental interactions, are the main targets, which will underpin the development of the crop for cultivation in an extensive range of environments. It is also needed to improve phytochemical content through traditional breeding programs or through bioengineering of the phenylpropanoid pathway, leading to desirable secondary metabolite accumulation. This paper is a review of modifications (in the field of biotechnology, agronomy, and processing) that have been applied to improve the nutritional value of B. napus L. Practical Applications: This paper has value for people starting their scientific work in the field related to the B. napus biotechnology and nutritional value of vegetable oils. In addition, it is a current compedium of knowledge for people working professionally in the oil industry. Moreover, this article is a cross‐section of the current achievements of genetic engineering in improving the composition of rapeseed and indicates key publications on this subject. Brassica napus L., the third most important oil crop in the world after maize and oil palm, has a unique history among those crops that are targets of breeding and genetic modification. Effective tools of processing, biotechnology, and agronomy are essential to produce rapeseed oil with a high amount of bioactive compounds and continue to be employed in strengthening the product versatility. [ABSTRACT FROM AUTHOR]

Plant terpenoids are a large and highly diverse class of metabolites with an important role in the immune defense. They find wide industrial application as active pharmaceutical ingredients, aroma and fragrance compounds. Several Eremophila sp. derived terpenoids have been documented. To elucidate the terpenoid metabolism, the transcriptome of juvenile and mature Eremophila serrulata (A.DC.) Druce (Scrophulariaceae) leaves was sequenced and a transcript library was generated. We report on the first transcriptomic dataset of an Eremophila plant. IlluminaMiSeq sequencing (2 × 300 bp) revealed 7,093,266 paired reads, which could be assembled to 34,505 isogroups. To enable detection of terpene biosynthetic genes, leaves were separately treated with methyl jasmonate, a well-documented inducer of plant secondary metabolites. In total, 21 putative terpene synthase genes were detected in the transcriptome data. Two terpene synthase isoenzymatic genes, termed ES01 and ES02, were successfully expressed in E. coli. The resulting proteins catalyzed the conversion of geranyl pyrophosphate, the universal substrate of monoterpene synthases to myrcene and Z-(b)-ocimene, respectively. The transcriptomic data and the discovery of the first terpene synthases from Eremophila serrulata are the initial step for the understanding of the terpene metabolism in this medicinally important plant genus., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Australian members of the Myrtaceae produce large quantities of ecologically and economically important terpenes and display abundant diversity in both yield and composition of their oils. In a survey of the concentrations of leaf terpenes in Eucalyptus tricarpa (L.A.S. Johnson) L.A.S. Johnson & K.D. Hill, which were previously known from few samples, exceptional variability was found in composition. The aim was to characterize the patterns of variation and covariation among terpene components in this species and to use this information to enhance our understanding of their biosynthesis. There were marked discontinuities in the distributions of numerous compounds, including the overall proportions of mono- and sesquiterpenes, leading us to delineate three distinct chemotypes. Overall, positive covariation predominated, but negative covariation suggested competitive interactions involved in monoterpene synthesis. Two groups of covarying monoterpenes were found, each of which was positively correlated with a group of sesquiterpenes and negatively correlated with the alternate sesquiterpene group. These results imply substantial cross-talk between mono- and sesquiterpene biosynthesis pathways. However, only those compounds hypothesized to share final carbocation intermediates or post-processing steps were strongly positively correlated within chemotypes. This suggests that the broader patterns of covariation among groups of compounds may result from co-regulation of multiple biosynthetic genes, controlling the complex terpene profiles of the chemotypes of Eucalyptus., (Copyright © 2013 Elsevier Ltd. All rights reserved.)