Factors influencing the efficiency of T-DNA transfer during co-cultivation of Antirrhinum majus with Agrobacterium tumefaciens

Summary The effects of varying the pH of the cocultivation medium, additons of vir-inducing phenolic compounds and the strains of wild-type agrobacteria on transformation rates of a number of different varieties of Antirrhinum majus were studied. In general, optimal transformation was found with strains C58 or A281 and was favoured by low pH and the inclusion of acetosyringone in the co-cultivation medium. However, maximal transformation of the least susceptible variety was achieved at high pH and in the presence of syringaldehyde. This demonstrates the need for the optimization of a wide range of culture conditions when working with new genotypes and offers a rational approach towards the development of Agrobacterium-mediated transformation of new species or varieties.Abbreviations BAP 6-benzylaminopurine - MS Murashige and Skoog medium (Murashige and Skoog, 1962) - NOA naphthoxyacetic acid     

Accumulation of calcium in the centre of leaves of coriander (Coriandrum sativum L.) is due to an uncoupling of water and ion transport

The aim of this study is to understand the parameters regulatingcalcium ion distribution in leaves. Accumulation of ions inleaf tissue is in part dependent on import from the xylem. Thisimport via the transpiration stream is more important for ionssuch as calcium that are xylem but not phloem mobile and cannottherefore be retranslocated. Accumulation of calcium was measuredon bulk coriander leaf tissue (Coriandrum sativum L. cv. Lemon)using ion chromatography and calcium uptake was visualized usingphosphor-images of ⁴⁵Ca²⁺. Leaves of plants grown in hydroponicshad elevated calcium in the centre of the leaf compared withthe leaf margin, while K⁺ was distributed homogeneously overthe leaf. This calcium was shown to be localised to the mesophyllvacuoles using EDAX. Stomatal density and evapotranspiration(water loss per unit area of leaf) were equal at inner and outersections of the leaf. Unequal ion distribution but uniformityof water loss suggested that there was a difference in the extentof uncoupling of calcium and water transport between the innerand outer leaf. Since isolated tissue from the inner and outerleaf were able to accumulate similar amounts of calcium, itis proposed that the spatial variation of leaf calcium concentrationis due to differential ion delivery to the two regions ratherthan tissue/cell-specific differences in ion uptake capacity.There was a positive correlation between whole leaf calciumconcentration and the difference in calcium concentration betweeninner and outer leaf tissue. Exposing the plants to increasedhumidity reduced transpiration and calcium delivery to the leafand abolished this spatial variation of calcium concentration.Mechanisms of calcium delivery to leaves are discussed. An understandingof calcium delivery and distribution within coriander will informstrategies to reduce the incidence of calcium-related syndromessuch as tip-burn and provides a robust model for the transportof ions and other substances in the leaf xylem. Key words: Calcium, Coriandrum sativum, distribution, ion chromatography, leaves, radioisotope, spatial variation, transpiration, uptake Received 29 August 2008; Accepted 16 October 2008     

Development of simple sequence repeat (SSR) markers for the assessment of gene flow between sea beet (Beta vulgaris ssp. maritima) populations

Molecular markers can be used to estimate gene flow indirectly by monitoring the relative frequency of alleles in adjacent populations. Sea beet (Beta vulgaris ssp. maritima) is a wild plant species found along the coastlines of many European countries and is closely related to cultivated beets. A set of six simple sequence repeat (SSR) markers that are polymorphic in UK populations have been developed for sea beet to assess the problems of indirect measurement of gene flow in these populations.     

The effects of anther culture and plant genetic background on Agrobacterium rhizogenes-mediated transformation of commercial cultivars and derived doubled-haploid Brassica oleracea

The production of transgenic roots was scored for eight Brassica oleracea cultivars from broccoli, cabbage, cauliflower and kale following inoculation with an Agrobacterium rhizogenes cell line carrying a binary plasmid bearing the green fluorescence protein (gfp) gene in the T-DNA. Significant differences in the numbers of explants producing transgenic roots were observed between cultivars, ranging from 1.4% for Marathon F1 to 57.8% for the Green Duke F1. Three F1 cultivars were subjected to anther culture, and doubled-haploid (DH) lines were used for transformation. The DH lines produced showed considerable variation for transgenic root production with some lines showing increased efficiency compared to the parental F1 cultivar. Grouping of the DH lines into response classes with respect to transgenic root production allowed the development of potential genetic models to explain the variation in performance released from each F1 cultivar. No apparent segregation distortion for transgenic root production was observed in the DH lines following anther culture.     

Development of simple sequence repeat (SSR) markers for the assessment of gene flow and genetic diversity in pigeonpea (Cajanus cajan)

Pigeonpea (Cajanus cajan) is an important subsistence crop in India where traditional landraces and improved hybrids are grown alongside each other. Gene flow may result in genetic erosion of these landraces and their wild relatives, whilst transgene escape from future genetically engineered varieties is another potential hazard. To assess the impact of these factors gene flow needs to be measured. A set of 10 simple sequence repeat markers have been developed, which exhibit polymorphism across a range of pigeonpea varieties. Use of these markers also offers an efficient system for the assessment of genetic diversity within populations of pigeonpea.     

Structural studies of microsporogenesis in fertile and male-sterile onions (Allium cepa L.) containing the cms-S cytoplasm

Summary The structure of anther tissues has been studied during microsporogenesis in male-sterile and -fertile onions. Three types of abnormal tapetal behaviour have been observed within the single line II/3ms containing the cms-S cytoplasm: type 1, the premature breakdown of the tapetum at the tetrad stage, type 2, the hypertrophy of the tapetum after the diad stage followed by its premature autolysis and, type 3, in which the tapetum remains in good condition but for an abnormally long period of time. Tapetal autolysis proceeds in the same manner in both male-steriles and -fertiles with only the stage at which it occurs differing between the types of plants. Mitochondria were prominent in the tapetal tissue of all onion types throughout all stages of microsporogenesis and were still visible during the last stages of tapetal autolysis. In a detailed study of type 2 behaviour, no differences in mitochondrial volumes were found until the tapetum hypertrophied.     

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.