Quantitative and molecular genetics of juvenile wood traits in radiata and slash/Caribbean pines.

The Juvenile Wood Initiative (JWI) project has been running successfully since July 2003 under a Research Agreement with FWPA and Letters of Association with the consortium partners STBA (Southern Tree Breeding Association), ArborGen and FPQ (Forestry Plantations Queensland). Over the last five and half years, JWI scientists in CSIRO, FPQ, and STBA have completed all 12 major milestones and 28 component milestones according to the project schedule. We have made benchmark progress in understanding the genetic control of wood formation and interrelationships among wood traits. The project has made 15 primary scientific findings and several results have been adopted by industry as summarized below. This progress was detailed in 10 technical reports to funding organizations and industry clients. Team scientists produced 16 scientific manuscripts (8 published, 1 in press, 2 submitted, and several others in the process of submission) and 15 conference papers or presentations. Primary Scientific Findings. The 15 major scientific findings related to wood science, inheritance and the genetic basis of juvenile wood traits are: 1. An optimal method to predict stiffness of standing trees in slash/Caribbean pine is to combine gravimetric basic density from 12 mm increment cores with a standing tree prediction of MoE using a time of flight acoustic tool. This was the most accurate and cheapest way to rank trees for breeding selection for slash/Caribbean hybrid pine. This method was also recommended for radiata pine. 2. Wood density breeding values were predicted for the first time in the STBA breeding population using a large sample of 7,078 trees (increment cores) and it was estimated that selection of the best 250 trees for deployment will produce wood density gains of 12.4%. 3. Large genetic variation for a suite of wood quality traits including density, MFA, spiral grain, shrinkage, acoustic and non-acoustic stiffness (MoE) for clear wood and standing trees were observed. Genetic gains of between 8 and 49% were predicted for these wood quality traits with selection intensity between 1 to 10% for radiata pine. 4. Site had a major effect on juvenile-mature wood transition age and the effect of selective breeding for a shorter juvenile wood formation phase was only moderate (about 10% genetic gain with 10% selection intensity, equivalent to about 2 years reduction of juvenile wood). 5. The study found no usable site by genotype interactions for the wood quality traits of density, MFA and MoE for both radiata and slash/Caribbean pines, suggesting that assessment of wood properties on one or two sites will provide reliable estimates of the genetic worth of individuals for use in future breeding. 6. There were significant and sizable genotype by environment interactions between the mainland and Tasmanian regions and within Tasmania for DBH and branch size. 7. Strong genetic correlations between rings for density, MFA and MoE for both radiata and slash/Caribbean pines were observed. This suggests that selection for improved wood properties in the innermost rings would also result in improvement of wood properties in the subsequent rings, as well as improved average performance of the entire core. 8. Strong genetic correlations between pure species and hybrid performance for each of the wood quality traits were observed in the hybrid pines. Parental performance can be used to identify the hybrid families which are most likely to have superior juvenile wood properties of the slash/Caribbean F1 hybrid in southeast Queensland. 9. Large unfavourable genetic correlations between growth and wood quality traits were a prominent feature in radiata pine, indicating that overcoming this unfavourable genetic correlation will be a major technical issue in progressing radiata pine breeding. 10. The project created the first radiata pine 18 k cDNA microarray and generated 5,952 radiata pine xylogenesis expressed sequence tags (ESTs) which assembled into 3,304 unigenes. 11. A total of 348 genes were identified as preferentially expressed genes in earlywood or latewood while a total of 168 genes were identified as preferentially expressed genes in either juvenile or mature wood. 12. Juvenile earlywood has a distinct transcriptome relative to other stages of wood development. 13. Discovered rapid decay of linkage disequilibrium (LD) in radiata pine with LD decaying to approximately 50% within 1,700 base pairs (within a typical gene). A total of 913 SNPS from sequencing 177,380 base pairs were identified for association genetic studies. 14. 149 SNPs from 44 genes and 255 SNPs from a further 51 genes (total 95 genes) were selected for association analysis with 62 wood traits, and 30 SNPs were shortlisted for their significant association with variation of wood quality traits (density, MFA and MoE) with individual significant SNPs accounting for between 1.9 and 9.7% of the total genetic variation in traits. 15. Index selection using breeding objectives was the most profitable selection method for radiata pine, but in the long term it may not be the most effective in dealing with negative genetic correlations between wood volume and quality traits. A combination of economic and biological approaches may be needed to deal with the strong adverse correlation.

Location of major effect genes in sorghum (Sorghum bicolor (L.) Moench).

Major effect genes are often used for germplasm identification, for diversity analyses and as selection targets in breeding. To date, only a few morphological characters have been mapped as major effect genes across a range of genetic linkage maps based on different types of molecular markers in sorghum (Sorghum bicolor (L.) Moench). This study aims to integrate all available previously mapped major effect genes onto a complete genome map, linked to the whole genome sequence, allowing sorghum breeders and researchers to link this information to QTL studies and to be aware of the consequences of selection for major genes. This provides new opportunities for breeders to take advantage of readily scorable morphological traits and to develop more effective breeding strategies. We also provide examples of the impact of selection for major effect genes on quantitative traits in sorghum. The concepts described in this paper have particular application to breeding programmes in developing countries where molecular markers are expensive or impossible to access.

Negligible evidence for regional genetic population structure for two shark species Rhizoprionodon acutus (Rüppell, 1837) and Sphyrna lewini (Griffith & Smith, 1834) with contrasting biology

Biodiversity of sharks in the tropical Indo-Pacific is high, but species-specific information to assist sustainable resource exploitation is scarce. The null hypothesis of population genetic homogeneity was tested for scalloped hammerhead shark (Sphyrna lewini, n = 237) and the milk shark (Rhizoprionodon acutus, n = 207) from northern and eastern Australia, using nuclear (S. lewini, eight microsatellite loci; R. acutus, six loci) and mitochondrial gene markers (873 base pairs of NADH dehydrogenase subunit 4). We were unable to reject genetic homogeneity for S. lewini, which was as expected based on previous studies of this species. Less expected were similar results for R. acutus, which is more benthic and less vagile than S. lewini. These features are probably driving the genetic break found between Australian and central Indonesian R. acutus (F-statistics; mtDNA, 0.751–0.903, respectively; microsatellite loci, 0.038–0.047 respectively). Our results support the spatially homogeneous monitoring and management plan for shark species in Queensland, Australia.

Towards better management of Australia’s shark fishery: genetic analyses reveal unexpected ratios of cryptic blacktip species Carcharhinus tilstoni and C. limbatus

The common blacktip shark (Carcharhinus limbatus) and the Australian blacktip shark (C. tilstoni) are morphologically similar species that co-occur in subtropical and tropical Australia. In striking contrast to what has been previously reported, we demonstrate that the common blacktip shark is not rare in northern Australia but occurs in approximately equal frequencies with the Australian blacktip shark. Management of shark resources in northern Australia needs to take account of this new information. Species identification was performed using nucleotide sequences of the control, NADH dehydrogenase subunit 4 (ND4) and cytochrome oxidase I (COI) regions in the mitochondrial genome. The proportion of overall genetic variation (FST) between the two species was small (0.042, P < 0.01) based on allele frequencies at five microsatellite loci. We confirm that a third blacktip species (C. amblyrhynchoides, graceful shark) is closely related to C. tilstoni and C. limbatus and can be distinguished from them on the basis of mtDNA sequences from two gene regions. The Australian blacktip shark (C. tilstoni) was not encountered among 20 samples from central Indonesia that were later confirmed to be common blacktip and graceful sharks. Fisheries regulators urgently need new information on life history, population structure and morphological characters for species identification of blacktip shark species in Australia.

IUCN classification zones concord with, but underestimate, the population genetic structure of the zebra shark Stegostoma fasciatum in the Indo-West Pacific

The Indo-West Pacific (IWP), from South Africa in the western Indian Ocean to the western Pacific Ocean, contains some of the most biologically diverse marine habitats on earth, including the greatest biodiversity of chondrichthyan fishes. The region encompasses various densities of human habitation leading to contrasts in the levels of exploitation experienced by chondrichthyans, which are targeted for local consumption and export. The demersal chondrichthyan, the zebra shark, Stegostoma fasciatum, is endemic to the IWP and has two current regional International Union for the Conservation of Nature (IUCN) Red List classifications that reflect differing levels of exploitation: ‘Least Concern’ and ‘Vulnerable’. In this study, we employed mitochondrial ND4 sequence data and 13 microsatellite loci to investigate the population genetic structure of 180 zebra sharks from 13 locations throughout the IWP to test the concordance of IUCN zones with demographic units that have conservation value. Mitochondrial and microsatellite data sets from samples collected throughout northern Australia and Southeast Asia concord with the regional IUCN classifications. However, we found evidence of genetic subdivision within these regions, including subdivision between locations connected by habitat suitable for migration. Furthermore, parametric FST analyses and Bayesian clustering analyses indicated that the primary genetic break within the IWP is not represented by the IUCN classifications but rather is congruent with the Indonesian throughflow current. Our findings indicate that recruitment to areas of high exploitation from nearby healthy populations in zebra sharks is likely to be minimal, and that severe localized depletions are predicted to occur in zebra shark populations throughout the IWP region.

The extent of population genetic subdivision differs among four co-distributed shark species in the Indo-Australian archipelago

Background: The territorial fishing zones of Australia and Indonesia are contiguous to the north of Australia in the Timor and Arafura Seas and in the Indian Ocean to the north of Christmas Island. The area surrounding the shared boundary consists of a variety of bio-diverse marine habitats including shallow continental shelf waters, oceanic trenches and numerous offshore islands. Both countries exploit a variety of fisheries species, including whaler (Carcharhinus spp.) and hammerhead sharks (Sphyrna spp.). Despite their differences in social and financial arrangements, the two countries are motivated to develop complementary co-management practices to achieve resource sustainability. An essential starting point is knowledge of the degree of population subdivision, and hence fisheries stock status, in exploited species. Results: Populations of four commercially harvested shark species (Carcharhinus obscurus, Carcharhinus sorrah, Prionace glauca, Sphyrna lewini) were sampled from northern Australia and central Indonesia. Neutral genetic markers (mitochondrial DNA control region sequence and allelic variation at co-dominant microsatellite loci) revealed genetic subdivision between Australian and Indonesian populations of C. sorrah. Further research is needed to address the possibility of genetic subdivision among C. obscurus populations. There was no evidence of genetic subdivision for P. glauca and S. lewini populations, but the sampling represented a relatively small part of their distributional range. For these species, more detailed analyses of population genetic structure is recommended in the future. Conclusion: Cooperative management between Australia and Indonesia is the best option at present for P. glauca and S. lewini, while C. sorrah and C. obscurus should be managed independently. On-going research on these and other exploited shark and ray species is strongly recommended. Biological and ecological similarity between species may not be a predictor of population genetic structure, so species-specific studies are recommended to provide new data to assist with sustainable fisheries management.

The application of DArTseq technology to pineapple

DArTseq technology is potentially the most appropriate system to discover hundreds of polymorphic genomic loci, scoring thousands of unique genomic-wide DNA fragments in one single experiment, without requiring existing DNA sequence information. The DArT complexity reduction approach in combination with Illumina short read sequencing (Hiseq2000) was applied. To test the application of DArTseq technology in pineapple, a reference population of 13 Ananas genotypes from primitive wild accessions to modern cultivars was used. In a comparison of 3 systems, the combination of restriction enzymes PstI and MseI performed the best producing 18,900 DArT markers and close to 20,000 SNPs. Based on these markers genetic relationships between the samples were identified and a dendrogram was generated. The topography of the tree corresponds with our understanding of the genetic relationships between the genotypes. Importantly, the replicated samples of all genotypes have a dissimilarity of close to 0.0 and occupy the same positions on the tree, confirming high reproducibility of the markers detected. Eventually it is planned that molecular markers will be identified that are associated with resistance to Phytophthora cinnamomi (Pc), the most economically important pathogen of pineapple in Australia, as genetic resistance is known to exist within the Ananas. Marker assisted selection can then be utilized in a pineapple breeding program to develop cultivars resistant to Pc.

ACIAR Improving Postrainy Sorghum Varieties Grain and Fodder in India

Sorghum grown in India in the post-rainy season (Rabi) relies on residual soil moisture, and the crop is commonly exposed to terminal drought stress. But there is a ready market for its high-quality grain and stover (used as fodder on dairy farms). Steps to improve productivity while maintaining quality offer an attractive opportunity for sorghum farmers to improve their incomes. Genetically improving the efficiency of using stored soil moisture is a prime target to maximise grain/stover production and quality of Rabi sorghum. This project aims to achieve this through the application of DNA sequences known as quantitative trait loci (QTLs). The project scientists will introduce marker-assisted introgression of stay-green QTLs into sorghum lines, enhancing both the quality and the quantity of grain/stover of postrainy sorghum. They will also use modelling to identify the key physiological traits involved in a higher, more stable yield across water-limited environments of India and Australia, and the key stay-green QTLs contributing to these traits. The publicly available QTL isolines lines developed in this project will be the basis of new varieties to be bred in a subsequent phase.

Development of pedigree mapping for wheat

The aim of this project is to build on the work done so far in applying pedigree mapping principals to northern adapted wheat pedigreed to identify robust QTL for quality traits.

DNA markers linked to the R(2) rust resistance gene in sunflower (Helianthus annuus L.) facilitate anticipatory breeding for this disease variant.

Pre-emptive breeding for host disease resistance is an effective strategy for combating and managing devastating incursions of plant pathogens. Comprehensive, long-term studies have revealed that virulence to the R (2) sunflower (Helianthus annuus L.) rust resistance gene in the line MC29 does not exist in the Australian rust (Puccinia helianthi) population. We report in this study the identification of molecular markers linked to this gene. The three simple sequence repeat (SSR) markers ORS795, ORS882, and ORS938 were linked in coupling to the gene, while the SSR marker ORS333 was linked in repulsion. Reliable selection for homozygous-resistant individuals was efficient when the three markers, ORS795, ORS882, and ORS333, were used in combination. Phenotyping for this resistance gene is not possible in Australia without introducing a quarantinable race of the pathogen. Therefore, the availability of reliable and heritable DNA-based markers will enable the efficient deployment of this gene, permitting a more effective strategy for generating sustainable commercial cultivars containing this rust resistance gene.

Rapid microsatellite marker development for African mahogany ( Khaya senegalensis, Meliaceae) using next-generation sequencing and assessment of its intra-specific genetic diversity.

Khaya senegalensis (African mahogany or dry-zone mahogany) is a high-value hardwood timber species with great potential for forest plantations in northern Australia. The species is distributed across the sub-Saharan belt from Senegal to Sudan and Uganda. Because of heavy exploitation and constraints on natural regeneration and sustainable planting, it is now classified as a vulnerable species. Here, we describe the development of microsatellite markers for K. senegalensis using next-generation sequencing to assess its intra-specific diversity across its natural range, which is a key for successful breeding programs and effective conservation management of the species. Next-generation sequencing yielded 93943 sequences with an average read length of 234bp. The assembled sequences contained 1030 simple sequence repeats, with primers designed for 522 microsatellite loci. Twenty-one microsatellite loci were tested with 11 showing reliable amplification and polymorphism in K. senegalensis. The 11 novel microsatellites, together with one previously published, were used to assess 73 accessions belonging to the Australian K. senegalensis domestication program, sampled from across the natural range of the species. STRUCTURE analysis shows two major clusters, one comprising mainly accessions from west Africa (Senegal to Benin) and the second based in the far eastern limits of the range in Sudan and Uganda. Higher levels of genetic diversity were found in material from western Africa. This suggests that new seed collections from this region may yield more diverse genotypes than those originating from Sudan and Uganda in eastern Africa.

Genetic effects and genetic relationships among shrunken (sh2) sweet corn lines and F1 hybrids

The objectives of this study were to quantify the components of genetic variance and the genetic effects, and to examine the genetic relationship of inbred lines extracted from various shrunken2(sh2) breeding populations. Ten diverse inbred lines developed from genetic background, were crossed in half diallel. Parents and their F1 hybrids were evaluated at three environments. The parents were genotyped using 20 polymorphic simple sequence repeats (SSR). Agronomic and quality traits were analysed by a mixed linear model according to additive-dominance genetic model. Genetic effects were estimated using an adjusted unbiased prediction method. Additive variance was more important than dominance variance in the expression of traits related to ear aspects (husk ratio and percentage of ear filled) and eating quality (flavour and total soluble solids). For agronomic traits, however, dominance variance was more important than additive variance. The additive genetic correlation between flavour and tenderness was strong (r = 0.84, P <0.01). Flavour, tenderness and kernel colour additive genetic effects were not correlated with yield related traits. Genetic distance (GD), estimated from SSR profiles on the basis of Jaccard's similarity coefficient varied from 0.10 to 0.77 with an average of 0.56. Cluster analysis classified parents according to their pedigree relationships. In most studied traits, F1 performance was not associated with GD.

Raw data for "Linkage disequilibrium estimation of effective population size with immigrants from divergent populations: a case study on Spanish mackerel (Scomberomorus commerson)."

Abstract of Macbeth, G. M., Broderick, D., Buckworth, R. & Ovenden, J. R. (In press, Feb 2013). Linkage disequilibrium estimation of effective population size with immigrants from divergent populations: a case study on Spanish mackerel (Scomberomorus commerson). G3: Genes, Genomes and Genetics. Estimates of genetic effective population size (Ne) using molecular markers are a potentially useful tool for the management of endangered through to commercial species. But, pitfalls are predicted when the effective size is large, as estimates require large numbers of samples from wild populations for statistical validity. Our simulations showed that linkage disequilibrium estimates of Ne up to 10,000 with finite confidence limits can be achieved with sample sizes around 5000. This was deduced from empirical allele frequencies of seven polymorphic microsatellite loci in a commercially harvested fisheries species, the narrow barred Spanish mackerel (Scomberomorus commerson). As expected, the smallest standard deviation of Ne estimates occurred when low frequency alleles were excluded. Additional simulations indicated that the linkage disequilibrium method was sensitive to small numbers of genotypes from cryptic species or conspecific immigrants. A correspondence analysis algorithm was developed to detect and remove outlier genotypes that could possibly be inadvertently sampled from cryptic species or non-breeding immigrants from genetically separate populations. Simulations demonstrated the value of this approach in Spanish mackerel data. When putative immigrants were removed from the empirical data, 95% of the Ne estimates from jacknife resampling were above 24,000.

Soral synapomorphies are significant for the systematics of the Ustilago-Sporisorium-Macalpinomyces complex (Ustilaginaceae)

The genera Ustilago, Sporisorium and Macalpinomyces are a polyphyletic complex of plant pathogenic fungi. The four main morphological characters used to define these genera have been considered homoplasious and not useful for resolving the complex. This study re-evaluates character homology and discusses the use of these characters for defining monophyletic groups recovered from a reconstructed phylogeny using four nuclear loci. Generic delimitation of smut fungi based on their hosts is also discussed as a means for identifying genera within this group. Morphological characters and host specificity can be used to circumscribe genera within the Ustilago-Sporisorium-Macalpinomyces complex.

QTL for root angle and number in a population developed from bread wheats (Triticum aestivum) with contrasting adaptation to water-limited environments

Root architecture traits in wheat are important in deep soil moisture acquisition and may be used to improve adaptation to water-limited environments. The genetic architecture of two root traits, seminal root angle and seminal root number, were investigated using a doubled haploid population derived from SeriM82 and Hartog. Multiple novel quantitative trait loci (QTL) were identified, each one having a modest effect. For seminal root angle, four QTL (-log10(P) >3) were identified on 2A, 3D, 6A and 6B, and two suggestive QTL (-log10(P) >2) on 5D and 6B. For root number, two QTL were identified on 4A and 6A with four suggestive QTL on 1B, 3A, 3B and 4A. QTL for root angle and root number did not co-locate. Transgressive segregation was found for both traits. Known major height and phenology loci appear to have little effect on root angle and number. Presence or absence of the T1BL.1RS translocation did not significantly influence root angle. Broad sense heritability (h 2) was estimated as 50 % for root angle and 31 % for root number. Root angle QTL were found to be segregating between wheat cultivars adapted to the target production region indicating potential to select for root angle in breeding programs. © 2013 Springer-Verlag Berlin Heidelberg.