B. V. Ford-Lloyd

Use of RAPD for the study of diversity within plant germplasm collections

As part of the development of a molecular toolkit for the study of diversity within large plant germplasm collections, RAPD technology has been applied to accessions of rice (Oryza sativa) obtained from the major world collection held at IRRI (the International Rice Research Institute) which supplies germplasm to breeders. Methods for the speedy extraction of DNA representative of a rice accession, its amplification by PCR to reveal reproducible products, and the analysis of the banding data using numerical techniques have been established. The biological meaningfulness of RAPD data has also been demonstrated by reference to previous work on classification and crossability.

The effects of acetosyringone and pH on Agrobacterium-mediated transformation vary according to plant species

SummaryExpiants of five plant species (Allium cepa, Antirrhinum majus, Brassica campestris. Glycine max, and Nicotiana tabacum) were co-cultivated with three Agrobacterium tumefaciens strains under different conditions to assess the effects of acetosyringone and medium pH on strain virulence. Tumours were incited on all dicotyledonous species by strains N2/73 and A281. The presence of acetosyringone during co-cultivation generally enhanced the virulence of these strains, most markedly N2/73 on A. majus and G. max, and A281 on G. max. Strain Ach5 was virulent only on N. tabacum in the absence of acetosyringone, which, when present, extended the host range to include A. majus. There was evidence to suggest that acetosyringone may suppress virulence in some strain/plant species interactions. Virulence was affected in some cases by medium pH, but there was no general effect across plant species.

Detection of somaclonal variation in garlic (Allium sativum L.) using RAPD and cytological analysis

Abstract Plants were regenerated by somatic embryogenesis from long-term callus cultures derived from five garlic (Allium sativum L.) cultivars. Thirty-five of these plants were subjected to RAPD analysis. The frequency of variation was found to be cultivar dependent: approximately 1% in the two clones Solent White and California Late and around 0.35% in another three clones, Chinese, Long Keeper and Madena. Certain band changes were found in regenerants of different cultivars, suggesting the existence of a mutation-sensitive part of the garlic genome. The karyotypes of another 75 regenerants derived from the same callus cultures of three parental garlic clones were examined. Of these plants, 9.3% were found to be tetraploids, 4% aneuploid and 2.6% showed a change in the position of the secondary constriction. No association could be shown between the rate of variation for molecular and cytological characters either by comparing cultivars or examining individual regenerants.

Predicting quantitative variation within rice germplasm using molecular markers

Diverse Asian rice (Oryza sativa) germplasm has been used to identify associations between various quantitative traits and RAPD molecular markers using multiple regression analysis. This has allowed us to predict for other samples of germplasm their performance for traits such as culm length and number, days to flowering, grain width, and panicle and leaf length using only RAPD marker data. Such predictive capability is possible because of the availability of extensive diversity held in genebanks, and can be used in the future to facilitate the exploitation of that biodiversity. More specifically the methodology could facilitate crop improvement by rapid ideotype prediction. For the mapping and isolation of QTLs (genes controlling quantitative traits) the method would provide information to guide the selection of parental material for hybridization and markers expected to show linkage to QTLs. It may also be possible that these associations could lead the way towards marker-assisted selection during breeding programs. In the future, this demonstration of association between markers and easily measured traits could also be extended to the study of important adaptive traits, such as stress tolerance, found either within germplasm collections or in natural populations.

Contrasting genetic diversity relationships are revealed in rice ( Oryza sativa L.) using different marker types

Genetic variation between samples of Oryza sativa from 19 localities in Bangladesh and Bhutan was assessed using two PCR-based molecular marker systems: RAPD (random amplification of polymorphic DNA) and ISSR-PCR (inter-simple sequence repeat polymerase chain reaction). Employing RAPD, a set of 14 decanucleotides of arbitrary sequence directed the amplification of 94 reproducible marker bands, 47 (50%) of which were polymorphic. In addition, a set of 9 ISSR primers were used to direct amplification of 71 PCR products, 40 (56%) of which were polymorphic. Multivariate analyses of the two PCR-based molecular marker data sets provided evidence that the patterns of variation correspond with the classification described by Glaszmann [9] using isozyme analysis. Subtle differences in the relationships revealed between rice groups using the two types of PCR-based marker led to investigations of their map positions using an intraspecific doubled haploid mapping population. The observation that the chromosomal locations of markers can influence diversity assessments is presented and the significance of this is discussed.

Complementary conservation strategies

The challenge facing the world’s biological and conservation scientists is threefold: to classify the existing biological diversity; to halt the rate of ecosystem, habitat, species and genetic loss; and to feed the ever increasing human population. It is generally agreed that a catastrophic loss of plant genetic diversity is occurring at this moment: species, gene combinations and alleles are being lost for ever and this process of genetic erosion is likely to become even more grave in the future. The conservation of plant diversity is of critical importance, because of the direct benefits to humans that can arise from its exploitation in new agricultural and horticultural crops, the development of medicinal drugs and the pivotal role played by plants in the functioning of all natural ecosystems. The economic, political and social consequences that would result from a steady loss of plant diversity combined with rapid population growth is likely to be devastating if unchecked. The importance of these issues to humankind is underlined in Article 1 of the objectives of the Convention on Biological Diversity (UNCED, 1992): The objectives of this convention ... are the conservation of biological diversity, the sustainable use of its components and the fair and equitable sharing of the benefits arising out of the utilisation of genetic resources ...

The identification of duplicate accessions within a rice germplasm collection using RAPD analysis

A set of accessions of Oryza sativa from the International Rice Research Institute (Philippines) that included known and suspected duplicates as well as closely related germplasm has been subjected to RAPD analysis. The number of primers, the number of polymorphic bands and the total number of bands were determined that will allow the accurate discrimination of these categories of accessions, including the identification of true and suspected duplicates. Two procedures have been described that could be employed on a more general basis for identifying duplicates in genetic resources collections, and further discussion on the values of such activities is presented.

Towards a definition of a crop wild relative

Crop wild relatives are an important socio-economic resource that is currently being eroded or even extinguished through careless human activities. If the Conference of the Parties (COP) to the CBD 2010 Biodiversity Target of achieving a significant reduction in the current rate of loss is to be achieved, we must first define what crop wild relatives are and how their conservation might be prioritised. A definition of a crop wild relative is proposed and illustrated in the light of previous Gene Pool concept theory. Where crossing and genetic diversity information is unavailable, the Taxon Group concept is introduced to assist recognition of the degree of crop wild relative relatedness by using the existing taxonomic hierarchy.

Effectiveness of different classes of molecular marker for classifying and revealing variation in rice (Oryza sativa) germplasm

We have examined the effectiveness of similar numbers of markers from four molecular marker systems (AFLP, isozymes, ISSR and RAPD) for revealing genetic diversity and discriminating between infraspecific groups of Oryza sativa germplasm. Each marker system classifies the germplasm into three major groups (most effectively with isozymes and AFLPs), but with differences (primarily with ISSR) between the precise classifications generated. However, at the highest levels of genetic similarity there was only partial agreement as to relationships between individual accessions when different markers were used. When variance was partitioned among and within the three subspecific groups, although the differences were not significant, greater variation was found among than within groups using AFLP and isozymes, with the reverse for RAPD and ISSR. Measurement of polymorphism using average heterozygosity and effective number of alleles gave similar results for each marker system. These results are discussed in relation to various genetic resources conservation activities, and the advisability of extrapolating to other sets of germplasm particularly of other crop species.

The Ex Situ conservation of plant genetic resources

Preface. Acronyms and Abbreviations. Figures, Plates, Tables and Appendices. 1. The Genetic Resources of Plants and Their Value to Mankind. 2. Evolution of Plants Under Domestication. 3. The Plant Genetic Resources Conservation Movement. 4. Preparing to Collect for ex situ Conservation. 5. Exploration and Field Collection. 6. Seed Gene Bank Conservation. 7. Field Gene Banks, Botanic Gardens in vitro, DNA and Pollen Conservation. 8. World ex situ Collections of Germplasm. 9. Community-Based Conservation. 10. Plant Genetic Resource Utilization. 11. Genetic Conservation Information Management. 12. Conservation Case Studies. 13. The Future of ex situ Conservation. References. Appendices. Index.