Megan E Jones

Development of robust PCR-based DNA markers for each homoeo-allele of granule-bound starch synthase and their application in wheat breeding programs

The absence of expression of the granule-bound starch synthase I (GBSSI) allele from chromosome 4A of wheat is associated with improved starch quality for making Udon noodles. Several PCR-based methods for the analysis of GBSS alleles have been developed for application in wheat. A widely applied approach has involved a simple PCR followed by electrophoretic separation of DNA products on agarose gels. The PCR amplifies one band from each of the loci on chromosomes 4A (Wx-B1), 7A (Wx-A1), and 7D (Wx-D1), and the band from the Wx-B1 locus is diagnostic for the occurrence of the null Wx-B1 allele that is associated with improved starch quality. The reliable detection of the null Wx-B1 allele has been important in identifying wheat breeding lines. Allele-specific PCR has also been used to successfully detect the occurrence of the null Wx-B1 allele. In the present paper the various protocols were evaluated by testing a segregating double haploid population from a cross between Cranbrook and Halberd and the tests gave good agreement in different laboratories. The application of the DNAbased tests applied in wheat breeding programs provides one of the first examples of a molecular marker selection for a grain quality trait being successfully applied in an Australian wheat breeding program.

High-throughput DNA extraction from forest trees

It is difficult to extract pure high-quality DNA from trees, which may not be amenable to advances in extraction methods suitable for other plants. A new commercial high-throughput DNA extraction system, using a silica binding matrix for purification and a multisample mixer mill for tissue disruption, was evaluated for its suitability withEucalyptus spp.,Pinus spp., andAraucaria cunninghamii (hoop pine). DNA suitable for a range of molecular biology applications was successfully extracted from all genera. The method was highly reliable when tested in more than 500 preparations and could be adapted to different tree species with relatively minor modifications.

Chloroplast DNA variation and population structure in the widespread forest tree, Eucalyptus grandis

Recognition of genetic structure of populations and the ability to identify vulnerable populations is useful for the formation of conservation management strategies for plants. Eucalyptus grandis is a tall forest tree that has a major area of occurrence in subtropical eastern Australia, with smaller populations located in the east coast tropics. Many widespread forest species exhibit population differentiation that corresponds to geographic regions. However, Eucalyptus grandis appears to be an exception based on isozyme and morphological data. This is intriguing given a large discontinuity between northern populations and those in the southern part of the species range. In this study, the distribution of a maternally inherited chloroplast locus was examined because it was more likely to reveal genetic structure due to the slower evolution of the chloroplast genome and limited dispersal of seed in eucalypts. As expected, the GST for chloroplast DNA was higher than that for nuclear DNA but indicated low population differentiation for a forest tree species. Phylogeographic analysis indicated that the 15 populations grouped into three broad geographical regions; however, overall population structure was weak suggesting that the large geographical disjunction in the distribution of E. grandis may be relatively recent. A paradigm for conservation management of E. grandis based on chloroplast DNA haplotype distribution would take into account the low differentiation among populations.

A method for assessing arbuscular mycorrhizal fungi group distribution in tree roots by intergenic transcribed sequence variation

We identified five taxonomic groups of arbuscular mycorrhizal fungi (AMF) inside roots of young trees of six species of legumes and six species of non-legumes from a field site in southern Costa Rica using an AMF group-specific PCR assay of the intergenic transcribed sequence and 18S rRNA gene fragment. Assay specificity was verified by cloning and sequencing representatives from four of the five AMF groups. We found no difference in overall AMF diversity levels between legumes and non-legumes or between plant species. Some groups of AMF may associate more frequently with legumes than others, as Glomus Group A (Glomus mosseae/intradices group) representatives were detected more frequently in legumes than non-legumes relative to Glomus Group B (Glomus etunicatum/claroideum) representatives.

Molecular Markers in Tree Improvement: Characterisation and Use in Eucalyptus

Eucalypts are the most widely planted hardwood trees in the world, occupying a global estate of around 12million ha (Turnbull 1999). The genus comprises over 700 species, most of which are endemic to Australia, and its diverse membership offers species with adaptability to a range of exotic tropical and temperate conditions with high growth rates on productive sites (Eldridge et al. 1994). They are a major source of wood for paper pulp and construction timber, as well as fuelwood for industrial and domestic purposes in many developing countries (Eldridge et al. 1994). Domestication of eucalypts is still at an early stage, with most breeding populations only several generations removed from wild populations (Eldridge et al. 1994). The challenge for eucalypt breeders, as with most tree crops, is to make genetic gains in the face of long generations and delays in selecting mature traits, which can be as long as 20–30 years for wood properties. Eucalypts, having mixed mating systems, are predominantly outcrossing (rates between 0.7 and 0.92), and are thought to possess high levels of genetic load and exhibit deleterious effects when inbred (Eldridge 1970; Potts and Reid 1990; Myburg et al. 2000). Consequently, breeders tend to avoid inbreeding, instead they manage broadly based breeding as well as specialty populations to select for genetic gains (Eldridge et al. 1994). Eucalypts, as a group, are recognised as being promiscuous, with weak reproductive barriers amongst taxa (Pryor 1976), and a frequency of natural and artificial hybridisation that declines as taxonomic distance between parents increases (Griffin et al. 1988; Potts et al. 2001). Interspecific F1 hybrids feature in a number of breeding programs, often because they combine desirable characteristics from the parental taxa, but also for hybrid superiority imparted through heterosis, epistasis or trait complementarity (Nikles and Griffin 1992). Many tropical eucalypts are amenable to vegetative propagation, hence, one strategy for improvement is intensive within family selection followed by mass clonal reproduction of elite hybrid trees (Eldridge et al. 1994).