Genotypic variation among a diverse collection of red clover (Trifolium pratense L.) germplasm and in-vitro techniques for screening resistance to Sclerotinia trifoliorum Erikks in red clover : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Plant Breeding at Massey University, Palmerston North, New Zealand

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Red clover (Trifolium pratense L.) is the world’s second-ranking most important forage species after alfalfa (Medicago sativa L.) based on seed volumes produced, marketed and cultivar availability. In New Zealand, red clover account for 23% of legume sales by volume after white clover (62%) and contributes significantly to primary industries by providing high quality feed for livestock. However, production is currently constrained by several limiting factors. In order to address some of these limitations, there is a need to broaden the genetic base of existing cultivars through breeding by introducing sources of genetic diversity from germplasm representing different geographical regions. In the study presented, two major experiments were carried out aiming to investigate: 1. The genetic diversity among 40 selected world source of red clover germplasm together with 3 local cultivars for ten important morphological traits under field condition across three seasons. Univariate and multivariate analysis were used to estimate genotypic variation for each trait and assess inter-relationships for a range of traits respectively so as to identify distinct germplasm accessions based on seasonal morphological measurements. Results from variance component analysis indicate significant genotypic variation as well as moderate to high repeatability among the yield related traits. This indicates potential underlying additive genetic variation among the 40 germplasm accessions for yield related morphological traits. 2. The response of eleven selected New Zealand commercial cultivars to clover rot disease (Sclerotinia trifoliorum Erikss) through artificial inoculation under high disease pressure in glasshouse to identify source of resistance for further breeding purposes. In order to facilitate the glasshouse artificial inoculation and identify the sources of resistance, this experiment used the in-vitro culturing procedures for production of S. trifoliorum ascospores to inoculate red clover plants using two locally sourced S. trifoliorum isolates. Although in-vitro production of sclerotia has been successful, our attempt to produce ascospores was not possible leading to delay in artificial inoculation of plants in glasshouse. This result suggests the need to improve on in-vitro techniques for ascospore production, understand appropriate culture condition and determine right choice of S. trifoliorum isolate to facilitate fertilization. Generally, both experiments under this study has provided valuable source of information and identify potential untapped germplasm material for future prospects of red clover breeding to develop new cultivars with improved yield, persistence and other desirable traits suitable for New Zealand condition.