Simon P. Whittock

QTL influencing growth and wood properties in Eucalyptus globulus.

Regions of the genome affecting physical and chemical wood properties (quantitative trait loci (QTL)), as well as growth, were identified using a clonally replicated, outbred F2 family (112 genotypes, each with two ramets) of Eucalyptus globulus, planted in a field trial in north-west Tasmania. Traits studied were growth (assessed by stem diameter), wood density, cellulose content, pulp yield and lignin content. These traits are important in breeding for pulpwood, and will be important in breeding for carbon sequestration and biofuel production. Between one and four QTL were located for each trait, with each QTL explaining between 4% and 12% of the phenotypic variation. Several QTL for chemical wood properties were co-located, consistent with their high phenotypic correlations, and may reflect pleiotropic effects of the same genes. In contrast, QTL for density and lignin content with overlapping confidence intervals were considered to be due to independent genes, since the QTL effects were inherited from different parents. The inclusion of fully informative microsatellites on the linkage map allowed the determination of homology at the linkage group level between QTL and candidate genes in different pedigrees of E. globulus and different eucalypt species. None of the candidate genes mapped in comparable studies co-located with our major QTL for wood chemical properties, arguing that there are important candidate genes yet to be discovered.

Genetic control of coppice and lignotuber development in Eucalyptus globulus

The economics of short-rotation pulpwood plantations of Eucalyptus globulus as a coppice crop are influenced by stump survival and subsequent coppice growth rates. This study revealed significant genetic diversity in coppicing traits, both within and between subraces, following felling in a progeny trial after 9 years of growth. A total of 67% of trees coppiced after 14 months, but subraces varied from 43 to 73%. Heritabilities for coppice success (0.07) and subsequent growth (0.16–0.17) were low but statistically significant. Strong genetic correlation between presence/absence of coppice, the number of stems coppicing from the stump and modal coppice height, indicate that selection is possible by using the binary trait. The ability of a tree to coppice was genetically correlated with tree growth prior to felling (rg = 0.61) and with nursery-grown seedling traits, where large genetic differences were observed in the development of lignotubers. Coppicing was genetically correlated with the number of nodes with lignotubers (rg = 0.66) and seedling stem diameter at the cotyledonary node (rg = 0.91). These traits were uncorrelated with later age growth and with each other. The results suggest that coppicing is influenced by three independent mechanisms—lignotuber development, enlargement of the seedling stem at the cotyledonary node and vigorous growth—which enhance ability to survive catastrophic damage, and indicate that both lignotuber and coppice development can be altered by both natural and artificial selection.

High-throughput genotyping of hop ( Humulus lupulus L.) utilising diversity arrays technology (DArT)

Implementation of molecular methods in hop (Humulus lupulus L.) breeding is dependent on the availability of sizeable numbers of polymorphic markers and a comprehensive understanding of genetic variation. However, use of molecular marker technology is limited due to expense, time inefficiency, laborious methodology and dependence on DNA sequence information. Diversity arrays technology (DArT) is a high-throughput cost-effective method for the discovery of large numbers of quality polymorphic markers without reliance on DNA sequence information. This study is the first to utilise DArT for hop genotyping, identifying 730 polymorphic markers from 92 hop accessions. The marker quality was high and similar to the quality of DArT markers previously generated for other species; although percentage polymorphism and polymorphism information content (PIC) were lower than in previous studies deploying other marker systems in hop. Genetic relationships in hop illustrated by DArT in this study coincide with knowledge generated using alternate methods. Several statistical analyses separated the hop accessions into genetically differentiated North American and European groupings, with hybrids between the two groups clearly distinguishable. Levels of genetic diversity were similar in the North American and European groups, but higher in the hybrid group. The markers produced from this time and cost-efficient genotyping tool will be a valuable resource for numerous applications in hop breeding and genetics studies, such as mapping, marker-assisted selection, genetic identity testing, guidance in the maintenance of genetic diversity and the directed breeding of superior cultivars.

Quantitative trait loci in hop (Humulus lupulus L.) reveal complex genetic architecture underlying variation in sex, yield and cone chemistry

BackgroundHop (Humulus lupulus L.) is cultivated for its cones, the secondary metabolites of which contribute bitterness, flavour and aroma to beer. Molecular breeding methods, such as marker assisted selection (MAS), have great potential for improving the efficiency of hop breeding. The success of MAS is reliant on the identification of reliable marker-trait associations. This study used quantitative trait loci (QTL) analysis to identify marker-trait associations for hop, focusing on traits related to expediting plant sex identification, increasing yield capacity and improving bittering, flavour and aroma chemistry.ResultsQTL analysis was performed on two new linkage maps incorporating transferable Diversity Arrays Technology (DArT) markers. Sixty-three QTL were identified, influencing 36 of the 50 traits examined. A putative sex-linked marker was validated in a different pedigree, confirming the potential of this marker as a screening tool in hop breeding programs. An ontogenetically stable QTL was identified for the yield trait dry cone weight; and a QTL was identified for essential oil content, which verified the genetic basis for variation in secondary metabolite accumulation in hop cones. A total of 60 QTL were identified for 33 secondary metabolite traits. Of these, 51 were pleiotropic/linked, affecting a substantial number of secondary metabolites; nine were specific to individual secondary metabolites.ConclusionsPleiotropy and linkage, found for the first time to influence multiple hop secondary metabolites, have important implications for molecular selection methods. The selection of particular secondary metabolite profiles using pleiotropic/linked QTL will be challenging because of the difficulty of selecting for specific traits without adversely changing others. QTL specific to individual secondary metabolites, however, offer unequalled value to selection programs. In addition to their potential for selection, the QTL identified in this study advance our understanding of the genetic control of traits of current economic and breeding significance in hop and demonstrate the complex genetic architecture underlying variation in these traits. The linkage information obtained in this study, based on transferable markers, can be used to facilitate the validation of QTL, crucial to the success of MAS.

Varietal characterization of hop (Humulus lupulus L.) by GC-MS analysis of hop cone extracts

An approach is described for use in the varietal characterization of hop (Humulus lupulus L.) varieties. The study focuses on commercial hop varieties and was timed to coincide with the 2008 commercial hop harvest in Tasmania, Australia. Analysis of hop extracts was performed using GC-MS. A 60 m capillary column was employed to increase efficiency to permit the use of a quadrupole mass spectrometer in place of a time of flight mass spectrometer that is more commonly used for this type of analysis. A set of characterization functions were derived from discriminant analysis which were highly suitable for varietal characterization of the eight commercial varieties included in the study, namely Willamette, Victoria, Pride of Ringwood, Cascade, Southern Hallertau, Millennium, Southern Saaz, and Super Pride.

Integrating revenues from carbon sequestration into economic breeding objectives for Eucalyptus globulus pulpwood production

A system where carbon sequestration was directly dependent upon biomass production in a plantation was modelled to assess whether economic breeding objectives for the genetic improvement of Eucalyptus globulus were sensitive to potential revenues from carbon sequestration. Carbon dioxide equivalent accumulation in the biomass (CO2e) of the Australian E. globulus plantation estate established between 2004 and 2012 was estimated. Total carbon dioxide equivalent (CO2e) accumulation was in the order of ∼146 t CO2e ha−1, of which 62 t CO2e ha−1 were tradable in 2012 (the 1st Kyoto Protocol commitment period) and a further 30 t CO2e ha−1 were tradable in 2016 (a hypothetical second Kyoto protocol commitment period). The correlated response of breeding objectives with and without carbon revenues (ΔcGh1) never fell below 0.86 in sensitivity analysis, and the mean was 0.93. Where economic breeding objectives for the genetic improvement of Eucalyptus globulus for pulpwood plantations are based on maximizing net present value by increasing biomass production, the consideration of carbon revenues in economic breeding objectives will have no significant effect on the relative economic weights of the key economic traits, wood basic density and standing volume at harvest.RésuméUn système où la séquestration du carbone était directement dépendante de la production de biomasse en plantation a été modélisé pour déterminer si des objectifs d’amélioration économique pour l’amélioration génétique d’Eucalyptus globulus réagissaient sur des revenus potentiels à partir de la séquestration du carbone. Le dioxyde de carbone gaz équivalent de l’accumulation de biomasse (CO2e) par des plantations australiennes d’Eucalyptus globulus crées entre 2004 et 2012 a été estimé. L’accumulation de dioxyde de carbone (CO2e) était de l’ordre de ∼146 t CO2e ha−1, dont 62 t CO2e ha−1, étaient commercialisables en 2012 (période correspondant aux engagement du premier protocole de Kyoto) et 30 t CO2e ha−1 supplémentaires étaient commercialisables en 2016 (hypothétique second protocole de Kyoto). La réponse correspondant aux objectifs d’amélioration avec et sans recettes de carbone (ΔcGH1) n’est jamais tombée sous 0,86 en analyse de sensibilité, et la moyenne était 0,93. Là où les objectifs d’amélioration économique pour l’amélioration génétique d’Eucalyptus globulus pour des productions de pâte à papier étaient basées sur maximalisation de la valeur actuelle nette par un accroissement de la production de biomasse, la prise en compte des recettes du carbone dans des objectifs d’amélioration économique n’aura pas d’effets significatifs sur le poids économique relatif de ces traits économiques, densité de base du bois et volume de bois sur pied à la récolte.

Quantitative genetic parameters for yield, plant growth and cone chemical traits in hop (Humulus lupulus L.)

BackgroundMost traits targeted in the genetic improvement of hop are quantitative in nature. Improvement based on selection of these traits requires a comprehensive understanding of their inheritance. This study estimated quantitative genetic parameters for 20 traits related to three key objectives for the genetic improvement of hop: cone chemistry, cone yield and agronomic characteristics.ResultsSignificant heritable genetic variation was identified for α-acid and β-acid, as well as their components and relative proportions. Estimates of narrow-sense heritability for these traits (h2 = 0.15 to 0.29) were lower than those reported in previous hop studies, but were based on a broader suite of families (108 from European, North American and hybrid origins). Narrow-sense heritabilities are reported for hop growth traits for the first time (h2 = 0.04 to 0.20), relating to important agronomic characteristics such as emergence, height and lateral morphology. Cone chemistry and growth traits were significantly genetically correlated, such that families with more vigorous vegetative growth were associated with lower α-acid and β-acid levels. This trend may reflect the underlying population structure of founder genotypes (European and North American origins) as well as past selection in the Australian environment. Although male and female hop plants are thought to be indistinguishable until flowering, sex was found to influence variation in many growth traits, with male and female plants displaying differences in vegetative morphology from emergence to cone maturity.ConclusionsThis study reveals important insights into the genetic control of quantitative hop traits. The information gained will provide hop breeders with a greater understanding of the additive genetic factors which affect selection of cone chemistry, yield and agronomic characteristics in hop, aiding in the future development of improved cultivars.

Parallel comprehensive two-dimensional gas chromatography

We introduce an information rich analytical approach called parallel comprehensive two-dimensional gas chromatography (2GC×2GC). This parallel chromatography approach splits injected samples into two independent two-dimensional column ensembles and provides two GC×GC separations by using contra-directional thermal modulation. The first-dimension (1D) and second-dimension (2D) columns are connected using planar three-port microchannel devices, which are supplied with supplementary flow via two pressure controller modules. Precise carrier gas flow control at the junction of the 1D and 2D columns permits independent control of flow conditions in each separation column. The 2GC×2GC approach provides two entirely independent GC×GC separations for each injection. Analysis of hop (Humulus lupulus L.) essential oils is used to demonstrate the capability of the approach. The analytical performance of each GC×GC separation in the 2GC×2GC experiment is comparable to individual GC×GC separation with matching column configurations. The peak capacity of 2GC×2GC is about 2 times than that of single GC×GC system. The dual 2D chromatograms produced by this single detector system provide complementary separations and additional identification information by harnessing different selectivity provided by the four separation columns.

Molecular variation of hop mosaic virus isolates

Hop mosaic virus (HpMV), a member of the genus Carlavirus, is importance to hop production worldwide. We identified variation in nucleic and amino acid sequences among 23 HpMV isolates from Australia, the USA, the Czech Republic, South Africa and Japan using a 1,455-bp fragment covering the 3′ end of the virus genome including ORFs 4, 5 and 6. Three clusters of two or more isolates were identified in phylogenies of the total nucleotide sequence and the coat protein (ORF5) amino acid sequence. Two of these clusters combined in analyses of ORF4 and ORF6 amino acid sequences. Isolates from within and outside of Australia were found in each cluster, indicating that sequence variation was not associated with geographic source. Monitoring of HpMV variants in the field and evaluation of the impact of variants on vector association, rate of spread, and hop yield and quality can now be undertaken.

Chemotyping of new hop (Humulus lupulus L.) genotypes using comprehensive two-dimensional gas chromatography with quadrupole accurate mass time-of-flight mass spectrometry

Comprehensive two-dimensional gas chromatography with quadrupole accurate mass time-of-flight mass spectrometry (GC×GC-Q-TOFMS) is employed to profile Humulus lupulus L. (hop) essential oils. Comparison of characterised essential oils allows discrimination among chemotypes. Experimental and commercial hop genotypes displayed distinguishable chemotypic patterns among the volatile secondary metabolites making up their essential oils. In total, 210-306 unique compounds were detected (depending on specific genotype), with 99 of these compounds either positively or tentatively identified. Identified volatile secondary metabolites were grouped into esters, monoterpene hydrocarbons, oxygenated monoterpenes, sesquiterpene hydrocarbons, oxygenated sesquiterpenes and ketones. Terpenoids were the dominant chemical families across all hop genotypes analysed, representing between 67% and 90% of the total ion count. The multidimensional chromatographic profiles of hop essential oils are extremely information-rich, making GC×GC-Q-TOFMS useful for fast screening of new hybrid hop genotypes, and therefore informing breeding strategies to derive new commercial hop cultivars for the development of distinctive and desirable beers.