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De Lucas J.A.,La Trobe University | De Lucas J.A.,Molecular Plant Breeding Cooperative Research Center | Forster J.W.,La Trobe University | Forster J.W.,Molecular Plant Breeding Cooperative Research Center | And 5 more authors.
Crop and Pasture Science | Year: 2012

White clover is one of the most important pasture legumes in global temperate regions. It is an outcrossing, insect-pollinated species with gene flow occurring naturally between plants. A 2-year study was conducted to assess the relationship between gene flow and physical distance in white clover under field conditions in southern Australia. White clover plants exhibiting a red leaf mark phenotypic trait acted as pollen donors to recipient plants lacking leaf markings at distances up to 200m distant from the donor plants. Progeny were scored for the dominant red-leafed phenotype and gene flow was modelled. Paternity was confirmed using simple sequence repeat markers. A leptokurtic pattern of gene flow was observed under conditions designed to measure maximised gene flow with the majority of pollination occurring in the first 50m from the donor pollen source. The combined use of simple sequence repeat and visual markers confirmed that there was also a white clover pollen source in addition to the donor plants. This research confirms the difficulty in ensuring absolute containment of gene flow in an outcrossing species grown in an environment when endemic populations are known to exist. © CSIRO 2012.

Kaur S.,La Trobe University | Francki M.G.,Food Western Australia | Francki M.G.,Molecular Plant Breeding Cooperative Research Center | Francki M.G.,Murdoch University | And 3 more authors.
Plant Biotechnology Journal | Year: 2012

An understanding of nature and extent of nucleotide sequence variation is required for programmes of discovery and characterization of single nucleotide polymorphisms (SNPs), which provide the most versatile class of molecular genetic marker. A majority of higher plant species are polyploids, and allopolyploidy, because of hybrid formation between closely related taxa, is very common. Mutational variation may arise both between allelic (homologous) sequences within individual subgenomes and between homoeologous sequences among subgenomes, in addition to paralogous variation between duplicated gene copies. Successful SNP validation in allopolyploids depends on differentiation of the sequence variation classes. A number of biological factors influence the feasibility of discrimination, including degree of gene family complexity, inbreeding or outbreeding reproductive habit, and the level of knowledge concerning progenitor diploid species. In addition, developments in high-throughput DNA sequencing and associated computational analysis provide general solutions for the genetic analysis of allopolyploids. These issues are explored in the context of experience from a range of allopolyploid species, representing grain (wheat and canola), forage (pasture legumes and grasses), and horticultural (strawberry) crop. Following SNP discovery, detection in routine genotyping applications also presents challenges for allopolyploids. Strategies based on either design of subgenome-specific SNP assays through homoeolocus-targeted polymerase chain reaction (PCR) amplification, or detection of incremental changes in nucleotide variant dosage, are described. © 2011 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

Dracatos P.M.,Australian Department of Primary Industries and Fisheries | Dracatos P.M.,La Trobe University | Dracatos P.M.,Molecular Plant Breeding Cooperative Research Center | Keane P.J.,La Trobe University | And 3 more authors.
Australasian Plant Pathology | Year: 2010

Puccinia coronata (crown rust) is a damaging pathogen of ryegrasses used for both forage and turf applications. Alternative sources of crown rust pathogen resistance with enhanced durability are required for perennial ryegrass breeding programs. This approach may require pyramiding of major and minor resistance gene combinations into varieties, opposing the capacity of evolving pathogen populations to overcome complex resistance mechanisms. Crown rust pathogen populations have been homogenised into distinct lineages on detached leaves through the use of molecular genotyping technology. Subsequent propagation and inoculation of such spore lineages on defined host populations and individuals requires high inoculation efficiency, given the delivery of initially small spore quantities. In this study, an inoculation protocol was developed for effective uniform infection on susceptible host plant genotypes at both seedling and adult developmental stages. A randomised complete block design was used to test the protocol on glasshouse-grown plants from 10 elite perennial ryegrass cultivars. Significant quantitative differences in pustule number and latency period were detected between cultivars, suggesting that the protocol was highly effective at providing a uniform distribution of pustules in low and high urediniospore suspension volumes. The inoculation and quantification methods provide a framework for standardised assessment of crown rust infection under controlled environment conditions, and may in principle be applied to any Pucciniahost interaction. © 2010 Australasian Plant Pathology Society.

Louie G.V.,Salk Institute for Biological Studies | Bowman M.E.,Salk Institute for Biological Studies | Tu Y.,Australian Department of Primary Industries and Fisheries | Tu Y.,Molecular Plant Breeding Cooperative Research Center | And 8 more authors.
Plant Cell | Year: 2010

Lignin forms from the polymerization of phenylpropanoid-derived building blocks (the monolignols), whose modification through hydroxylation and O-methylation modulates the chemical and physical properties of the lignin polymer. The enzyme caffeic acid O-methyltransferase (COMT) is central to lignin biosynthesis. It is often targeted in attempts to engineer the lignin composition of transgenic plants for improved forage digestibility, pulping efficiency, or utility in biofuel production. Despite intensive investigation, the structural determinants of the regiospecificity and substrate selectivity of COMT remain poorly defined. Reported here are x-ray crystallographic structures of perennial ryegrass (Lolium perenne) COMT (Lp OMT1) in open conformational state, apo- and holoenzyme forms and, most significantly, in a closed conformational state complexed with the products S-adenosyl-L-homocysteine and sinapaldehyde. The product-bound complex reveals the postmethyl-transfer organization of COMT's catalytic groups with reactant molecules and the fully formed phenolic-ligand binding site. The core scaffold of the phenolic ligand forges a hydrogen-bonding network involving the 4-hydroxy group that anchors the aromatic ring and thereby permits only metahydroxyl groups to be positioned for transmethylation. While distal from the site of transmethylation, the propanoid tail substituent governs the kinetic preference of ryegrass COMT for aldehydes over alcohols and acids due to a single hydrogen bond donor for the C9 oxygenated moiety dictating the preference for an aldehyde. © American Society of Plant Biologists.

Dracatos P.M.,La Trobe University | Dracatos P.M.,Molecular Plant Breeding Cooperative Research Center | Cogan N.O.I.,La Trobe University | Keane P.J.,Molecular Plant Breeding Cooperative Research Center | And 6 more authors.
Crop Science | Year: 2010

The most serious foliar disease of perennial ryegrass (Lolium perenne L.) is crown rust (caused by Puccinia coronata Corda f.sp. lolii Brown). Progress in resistance breeding using recurrent selection has been slow, due to lack of genetic knowledge. Puccinia coronata is a basidiomycete fungus with a complex life-cycle involving both asexual and sexual reproductive modes. Pathotype variation may exist, but confi rmation is complicated by the outbreeding nature of the host grass and consequent intrapopulation diversity. Qualitative and quantitative resistance mechanisms may hence be due to either major resistance genes responding to mixed pathogen populations, or minor (quantitative) resistance genes. Advances in molecular biology have permitted analysis of both host and pathogen genetics. Development of pathogen-derived simple sequence repeat (SSR) genetic markers allowed detailed analysis of genetic variation, as well as generation of homogenized inoculum for detailed trait-dissection studies. Multiple genomic regions were identifi ed as containing quantitative trait loci (QTLs) for pathogen resistance, and specifi c single nucleotide polymorphism (SNP) markers for defense response (DR) and disease resistance (R) genes have been evaluated for QTL co-location. The current knowledge status of the crown rust pathogen and genetics of host resistance is reviewed, as well as future prospects for facilitated rust resistance breeding. © Crop Science Society of America.

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