Induction of high-frequency somatic embryos in cassava for cryopreservation

Methods for inducing high-frequency somatic embryos in cassava on cotyledons and 33 clonal accessions by the addition of supplementary copper sulphate to the induction medium were investigated. The addition of copper sulphate enhanced primary embryo induction and significantly increased secondary embryo production. All accessions from Latin America (CIAT) were embryogenically competent on medium supplemented with 8 mg l^-1 2,4-dichlorophenoxyacetic acid (2,4-D) plus 1 µM copper sulphate as were 15 of the 18 accessions from Africa. The percentage of calli producing somatic embryos ranged from 7.5% in M. Bra 12 to 100% in M. Col. 1505, while the number of embryos produced per callus ranged from 0.3 in M. Bra 383 to 13.5 in TEK. The frequency of embryo production was dependent on the concentration of copper sulphate. The number of primary embryos produced per callus was also comparatively higher in the medium supplemented with copper sulphate than in the controls. The optimal concentration of copper sulphate for number of embryos produced in most accessions was 5 µM, and at this concentration the number of embryos produced was double that of the controls. Copper sulphate also reduced the maturation time of somatic embryos to 25 days from embryo initiation. High levels of 2,4-D were detrimental to embryo production. Similarly, fragmented embryos incubated in the dark produced more embryos tan those incubated under light conditions. On the basis of these results, the use of cassava somatic embryo micropropagules for germplasm conservation and synthetic seed development seems to be a strong possibility.     

Encapsulation of nodal cuttings and shoot tips for storage and exchange of cassava germplasm

We report the encapsulation of in vitro-derived nodal cuttings or shoot tips of cassava in 3% calcium alginate for storage and germplasm exchange purposes. Shoot regrowth was not significantly affected by the concentration of sucrose in the alginate matrix while root formation was. In contrast, increasing the sucrose concentration in the calcium chloride polymerisation medium significantly reduced regrowth from encapsulated nodal cuttings of accession TME 60444. Supplementing the alginate matrix with increased concentrations of 6-benzylaminopurine and alpha-naphthaleneacetic acid enhanced complete plant regrowth within 2 weeks. Furthermore, plant regrowth by encapsulated nodal cuttings and shoot tips was significantly affected by the duration of the storage period as shoot recovery decreased from almost 100% to 73.3% for encapsulated nodal cuttings and 94.4% to 60% for shoot tips after 28 days of storage. The high frequency of plant regrowth from alginate-coated micropropagules coupled with high viability percentage after 28 days of storage is highly encouraging for the exchange of cassava genetic resources. Such encapsulated micropropagules could be used as an alternative to synthetic seeds derived from somatic embryos.     

Isolation of protoplasts and regeneration of callus from suspension cultures of cultivated beets

Conditions necessary for the isolation and culture of protoplasts from suspension cultures of sugar, fodder and garden beets were investigated. Good yields of protoplasts were obtained by treating cells with a mixture of cellulase, Macerozyme and Driselase enzymes. Nutritional requirements of beet protoplasts were found to be quite simple: protoplasts could be cultured in MS, B₅ or PGo based media with 0.4 M glucose with the optimum result being produced on KM8p medium. Plating efficiency (P.E) was genotype-dependent with the sugar beet giving better P.E. than the fodder or garden beets used, and higher values being achieved with the use of desalted Driselase for isolation followed by culture on KMBp medium.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - BAP N⁶ benzylaminopurine - NAA naphthaleneacetic acid - P.E. plating efficiency - * University of Birmingham beet germplasm accession number     

Isolation and partial amino acid sequencing of the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase from three species of orchid

Small subunits of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) have been purified from 3 species of orchid in the genus Cymbidium by gel filtration followed by preparative sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and electroelution. The samples were subjected to amino acid composition analysis and partial N-terminal amino acid sequencing. The sequencing data clearly confirm that one of the species examined is the hybrid offspring of a cross between the other two.     

Numerical analyses of RAPD data highlight the origin of cultivated tagasaste (Chamaecytisus proliferus ssp. palmensis) in the Canary Islands

Random Amplified Polymorphic DNA (RAPD) was used to generate molecular markers to trace the origin of the fodder legume tagasaste (Chamaecytisus proliferus (L. fil.) Link ssp. palmensis (H. Christ) Kunkel) in the Canary Islands. Results from multivariate analyses of data through Two Way Indicator Species Analysis (TWINSPAN) and Detrended Correspondence Analysis (DECORANA) showed that genotypes collected on the island of La Palma exhibited a wider range of variation than those from the other islands. This supports the existing hypothesis that tagasaste originated on La Palma and emphasizes the importance of conserving and evaluating germ plasm from this island.     

In vitro adventitious root induction in Antirrhinum majus L.

A broadly applicable method for the successful induction of root systems in a number of cultivars of A. majus has been determined. This involves a double filter-paper bridge with a liquid medium for root induction and allows the transfer of culture-grown plantlets to a glasshouse environment with minimal disturbance to the plant as a whole. 100% survival of transferred plantlets has been achieved with the inclusion of a few simple precautions upon shoot transfer and during the initial stages of plant establishment in vivo.     

A global approach to crop wild relative conservation: securing the gene pool for food and agriculture

In light of the growing concern over the potentially devastating impacts on biodiversity and food security of climate change and the massively growing world population, taking action to conserve crop wild relatives (CWR), is no longer an option — it is a priority. Crop wild relatives are species closely related to crops, including their progenitors, many of which have the potential to contribute beneficial traits to crops, such as pest or disease resistance, yield improvement or stability. They are a critical component of plant genetic resources for food and agriculture (PGRFA), have already made major contributions to crop production and are vital for future food security; their systematic conservation in ways that ensure their continuing availability for use is therefore imperative. This is a complex, interdisciplinary, global issue that has been addressed by various national and international initiatives. Drawing on the lessons learnt from these initiatives we can now propose a global approach to CWR conservation, the key elements of which are: (1) estimating global CWR numbers, (2) assessment of the global importance of CWR diversity, (3) current conservation status, (4) threats to CWR diversity, (5) systematic approaches to CWR conservation, (6) CWR informatics, and (7) enhancing the use of CWR diversity.     

Islands as refugia of <Emphasis Type="Italic">Trifolium repens</Emphasis> genetic diversity

Island populations are often thought to be more susceptible to the loss of genetic diversity as a consequence of limited population size and genetic drift, greater susceptibility to detrimental stochastic events and low levels of immigration. However the geographic isolation of islands may create refuges for native crop species whose genetic diversity is threatened from the genetic erosion occurring in mainland areas as a result of crop-wild gene flow and genetic swamping. Many UK islands remain uncharacterised in terms of plant genetic diversity. In this study we compared the genetic diversity of mainland populations and landraces of Trifolium repens with wild populations collected from the islands surrounding the UK, including the island of Hirta in the St Kildan archipelago. Individuals from St Kilda represent a unique conservation resource, with populations both highly differentiated from UK mainland populations and genetically distinct from cultivated varieties, whilst able to retain diversity through limited human influence on the islands. In contrast, there is relative genetic similarity of wild UK populations to cultivated forms highlighted in mainland populations, but with geographic barriers preventing complete homogenisation of the mainland UK genepool. We underline the need for conservation priorities to include common species that are threatened by gene flow from cultivation, and draw attention to the potential of islands to preserve natural levels of genetic diversity.     

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.     

Identification of Taxa in the Genus shape Beta using ITS1 Sequence Information

Sequence variation in the ITS1 locus of the nuclear ribosomal DNA in beets has previously been used to reconstruct phylogeny of the species in the genus Beta. We have developed protocols that allow the identification of Beta taxa by use of taxon-specific primers. Beta sections, species and subspecies can be identified. Differences within the ITS1 region of a single base can be exploited for species identification. The results from this study not only provide effective methods for wild beet identification, but also indicate the potential use of the techniques in other crops.