Ken H. Chorlton

An association mapping approach to identify flowering time genes in natural populations of Lolium perenne (L.)

We describe an association mapping approach using natural populations of perennial ryegrass (Lolium perenne L.) to identify molecular markers associated with heading date, an important trait affecting seasonal production, tillering, digestibility and grassland management regimes. Twenty-three natural populations originating from throughout Europe, with heading date phenotypes ranging from very early to very late, as well as three synthetic populations (varieties) were used for molecular marker genotyping using AFLP. In total, 589 polymorphic markers were identified. Hierarchical clustering, principal coordinate and other statistical analyses identified four outlying populations forming a clearly distinct sub-group. Removal of those four populations from the subsequent analysis reduced population sub-structure twofold. However, this made relatively little difference to the result of the association analysis. Linear regression identified three markers whose frequency of occurrence correlated with the heading date phenotype. Moreover, these markers were shown to be closely linked to each other within a major QTL on Chromosome 7, explaining 70% of the total variation in heading date. Pairwise linkage disequilibrium among them was also significant. These results suggest that association mapping approaches may be feasible in L. perenne, and that the use of natural populations could provide a useful source of genetic variation in traits of importance in crop improvement.

Molecular genecology of temperature response in Lolium perenne: 2. Association of AFLP markers with ecogeography

Improved winter hardiness is an important breeding objective in the forage grass Lolium perenne. This is a complex trait with several components, including the ability to survive and grow at low temperature, to acclimate to cold, tolerate wind, snow cover and ice encasement. Marker‐assisted selection has the potential to increase the efficiency of breeding for improved cold tolerance. Here we describe a genecological approach to identifying molecular markers that are associated with adaptation to low winter temperatures. AFLP was used to assess the genetic diversity in 29 wild populations of ryegrass (Lolium perenne) representing a pan‐European temperature cline in terms of their geographical origin. A further 18 populations from a temperature cline in Bulgaria were also analysed. In addition, two varieties and five populations representing parents of mapping families currently in use at IGER were included in the analysis. Principal coordinate (PCoA) and cluster analyses of the molecular marker data showed that the Bulgarian altitude cline populations could be distinguished clearly from the other populations. Two regression analyses were carried out; one to identify AFLP markers that correlated in frequency with low mean January temperature of the geographical origin of the population, and another to identify AFLP markers correlating in frequency with the cold tolerance phenotype of the populations, as determined by LT50 values in freezing tests. In the first analysis six AFLP markers showed significant type II trends with mean January temperature, and in the second analysis 28 bands had a significant univariate relationship with the LT50 value of the accessions. In steps 2 and 3 of the stepwise analysis a further 4 and 5 bands, respectively, improved the fit significantly. The results of the two types of regression analysis are discussed in relation to ecogeography and cold tolerance phenotype of the populations.

Molecular genecology of temperature response in Lolium perenne: 1. preliminary analysis to reduce false positives

Molecular genecology is the study of geographical clines in frequencies of molecular markers and their relationship to ecological clines in environmental conditions. This study outlines the principles underlying the selection of populations, focusing on avoiding ‘false positives’— noncausal correlations between allele frequency and the environment. The principles are illustrated by identifying a set of populations of Lolium perenne for the study of temperature responses. The selected set of populations encompasses a 20 °C range in mean January temperature. Their freezing tolerance shows a linear trend with winter temperature, LT50 decreasing by 0.25 °C for each 1 °C reduction in mean January temperature.

A forage grass and small grain legume plant collecting expedition in South East Poland, 1990

SummaryThe Plant Genetic Resources Unit (PGRU) of the Institute of Grassland and Environmental Research, Welsh Plant Breeding Station (IGER, WPBS), UK, and the Plant Genetic Resources Section of the Plant Breeding and Acclimatization Institute (PBAI), Poland, carried out a joint collecting expedition in Poland between 25 August and 7 September 1990. The expedition was unique in that it was the first time that vegetative sampling had been applied to the perennial forage grass and legume populations of south east Poland. The expedition focused on seminatural vegetation in agriculturally managed situations and detailed collection site data on management systems was obtained from landowners.Samples were collected from 62 sites. Vegetative collections of Lolium spp. (37 populations) and Trifolium spp. (56 populations), and seed collections of Festuca spp. (32 populations) were made by the IGER team (Table 2). The PBAI team made 59 separate seed collections, mainly of Leguminosae.The expedition covered four geographical subregions of south east Poland. (See Fig. 1). These were the Nizina Mazowiecka south of Warszawa, the Wyzyna Malopolska with the town of Kielce at its centre and the Beskidy Zachodnie and Beskidy Wschodnie regions of the Carpathian mountains.A diverse range of habitats was sampled covering a broad range of altitude agricultural management systems and ecological conditions. Polish agriculture is faced with the problems of a rapid orientation to a market economy and it is likely that the diverse range of habitats encoutered will be reduced as agricultural practices change. This will lead to genetic erosion of the unique forage grass and legume populations to be found in Poland.

A forage grass and legume plant collecting expedition in Czechoslovakia 1992

The Plant Genetic Resources Unit (PGRU) of the Institute of Grassland and Environmental Research, Welsh Plant Breeding Station (IGER, WPBS), UK, and the Grassland Research Station, ZubrŨí (GRS, Z), in 1992 part of OSEVA Research Institute for Fodder Crops, Troubsko (OSEVA, RIFC), Czechoslovakia, carried out a collaborative plant collecting expedition in Czechoslovakia between 10 August and 29 August 1992. 14 geographical sub-regions of Czechoslovakia were covered with centres of collection in Bohemia, Moravia and Slovakia. The expedition concentrated on a range of vegetation from managed semi-natural grassland to wild, un-managed grassland. The IGER team collected mainly vegetative samples and the GRS, Z team collected seed samples of forage grass and legume populations. Detailed collection site data was recorded at every site. Samples were collected from 67 sites. Vegetative collections of Lolium spp. (34 populations) and Trifolium spp. (39 populations) and seed collectio ns of other species (57 populations) were made by the IGER team. Seed collections of Lolium spp. (26 populations), Trifolium spp. (19 populations) and other species (28 populations) were made by the GRS, Z team (Table 2). A diverse range of habitats was sampled covering a range of altitude, management systems and ecological conditions. Wild grassland was easily found but semi-natural grassland was uncommon in the collective farming landscape. Entry into a free-market economy is forcing changes on the collective farming system which dominates Czechoslovakian agriculture. Changes in crops and management practices will lead to a further reduction in the area of semi-natural grassland and genetic erosion of the unique forage grass and legume populations in Czechoslovakia. The collecting expedition took place in August 1992, 5 months before Czechoslovakia was divided into the separate states of the Czech Republic and Slovakia. In the text ‘Czechoslovakia’ is used to reflect the situation at the time of the collecting expedition.