DNA based diagnostic for the quantification of sugarcane root DNA in the field.

Plant root systems play many key roles including nutrient and water uptake, interface with soil microorganisms and resistance to lodging. As for other crops, large and systematic studies of sugarcane root systems have always been hampered by the opaque and solid nature of the soil. In recent years, methods for efficient extraction of DNA from soil and for species-specific DNA amplification have been developed. Such tools could have potential to greatly improve root phenotyping and health diagnostic capability in sugarcane. In this paper, we present a fast, specific and efficient method for the quantification of sugarcane live root cells in soil samples. Previous studies were typically based on mass and length, so we established a calibration to convert root DNA quantity to live root mass. This diagnostic was validated on field samples and used to investigate the fate of the root system after harvest prior to regrowth of the ratoon crop. Two weeks after harvest, the sugarcane roots from the previous crop were still viable. This raises the question of the role that the root system of the harvested crop plays in the performance of the next crop and demonstrates how this test can be used to answer research questions.

Abiotic Limits for Germination of Sugarcane Seed in Relation to Environmental Spread.

Sugarcane is a vegetatively propagated crop and hence the production of seed and its fate in the environment has not been studied. The recent development of genetically modified sugarcane, with the aim of commercial production, requires a research effort to understand sugarcane reproductive biology. This study contributes to this understanding by defining the abiotic limits for sugarcane seed germination. Using seed from multiple genetic crosses, germination was measured under different light regimes (light and dark), temperatures (from 18 °C to 42 °C) and water potentials (from 0 MPa to -1 MPa); cardinal temperatures and base water potential of germination were estimated based on the rates of germination. We found that sugarcane seed could germinate over a broad range of temperatures (from 11 °C to 42 °C) with optima ranging from 27 °C to 36 °C depending on source of seed. Water potentials below -0.5 MPa halved the proportion of seed that germinated. By comparing these limits to the environmental conditions in areas where sugarcane grows and has the potential to produce seed, water, but not temperature, will be the main limiting factor for germination. This new information can be taken into account when evaluating any risk of weediness during the assessment of GM sugarcane.

Molecular mechanisms of phosphate and sulphate transport in plants.

The application of molecular techniques in recent years has advanced our understanding of phosphate and sulphate transport processes in plants. Genes encoding phosphate and sulphate transporters have been isolated from a number of plant species. The transporters encoded by these genes are related to the major facilitator superfamily of proteins. They are predicted to contain 12 membrane-spanning domains and function as H(+)/H(2)PO(-4) or H(+)/SO(2/-4) cotransporters. Both high-affinity and low-affinity types have been identified. Most research has concentrated on genes that encode transporters expressed in roots. The expression of many of these genes is transcriptionally regulated by signals that respond to the nutrient status of the plant. Nutrient demand and the availability of precursors needed in the assimilatory pathways also regulate transcription of some of these genes. Information on the cell types in which phosphate and sulphate transporters are expressed is becoming available. These data, together with functional characterisation of the transporters, are enabling the roles of various transporters in the overall phosphate and sulphate nutrition of plants to be defined.

Inhibition of transforming growth factor alpha (TGF-alpha)-mediated growth effects in ovarian cancer cell lines by a tyrosine kinase inhibitor ZM 252868.

The modulating effects of the epidermal growth factor (EGF) receptor-specific tyrosine kinase inhibitor ZM 252868 on cell growth and signalling have been evaluated in four ovarian carcinoma cell lines PE01, PE04, SKOV-3 and PE01CDDP. Transforming growth factor alpha (TGF-alpha)-stimulated growth was completely inhibited by concentrations > or =0.3 microM in the PE01 and PE04 cell lines and by > or =0.1 microM in SKOV-3 cells. TGF-alpha inhibition of PE01CDDP growth was reversed by concentrations > or =0.1 microM ZM 252868. TGF-alpha-stimulated tyrosine phosphorylation of both the EGF receptor and c-erbB2 receptor in all four cell lines. The inhibitor ZM 252868, at concentrations > or =0.3 microM, completely inhibited TGF-alpha-stimulated tyrosine phosphorylation of the EGF receptor and reduced phosphorylation of the c-erbB2 protein. EGF-activated EGF receptor tyrosine kinase activity was completely inhibited by 3 microM ZM 252868 in PE01, SKOV-3 and PE01CDDP cells. These data indicate that the EGF receptor-targeted TK inhibitor ZM 252868 can inhibit growth of ovarian carcinoma cells in vitro consistent with inhibition of tyrosine phosphorylation at the EGF receptor.

A peroxidase gene promoter induced by phytopathogens and methyl jasmonate in transgenic plants.

The expression of two closely related peroxidase isogenes, Shpx6a and Shpx6b, of the legume Stylosanthes humilis was studied using isogene-specific reverse transcriptase PCR techniques. Results indicated that transcripts of both genes were rapidly induced following inoculation with the fungal pathogen Colletotrichum gloeosporioides, wounding and treatment with the defense regulator methyl jasmonate (MeJA). In contrast treatment of leaves of S. humilis with abscisic acid (ABA) and salicylic acid (SA) did not induce transcripts of either isogene. A genomic clone containing the Shpx6b gene was isolated and 594 bp of 5' sequence upstream of the translation start was fused in frame to the coding region of the uidA reporter gene and introduced into tobacco. Expression from the Shpx6b promoter in transgenic plants was determined by histochemical staining and quantitative assays of beta-glucuronidase (GUS). In transgenic tobacco, GUS expression was detected in cotyledons, vascular cells of young leaves, anthers, pollen, and the stigma and style. Wounding of the tobacco plants produced very localized GUS staining. Much more extensive staining for GUS was observed following inoculation of tobacco leaves with conidia of the fungal pathogen Cercospora nicotianae and the inoculation of wound sites with mycelium of the Oomycete pathogen Phytophthora parasitica var. nicotianae. Treatment of mature leaves with methyl jasmonate induced GUS activity while treatment with ABA, SA, and H2O2 had no effect. A similar strong induction of GUS activity was measured in young transgenic seedlings germinated on MeJA while some, but much weaker, induction of GUS activity was observed in seedlings treated with SA. The sequence of the promoter contained motifs homologous to putative cis elements in other plant genes responsive to MeJA. The Shpx6b gene is the first plant peroxidase gene shown to be induced by both microbial pathogens and MeJA and its promoter will be useful for investigations of signaling processes during fungal infection and for the expression of foreign gene products at infection sites.

Bacterial and plant glycoconjugates at the Rhizobium-legume interface.

Many classes of bacterial and plant glycoconjugate have been shown to be involved in establishing the Rhizobium root nodule symbiosis with peas (Pisum sativum). It was demonstrated, using techniques of molecular genetics, that a group of Rhizobium nodulation genes (nod genes) co-operate to synthesize a lipo-oligosaccharide signal molecule that specifically initiates nodule development on legume hosts. An additional gene function, encoded by nodX, has been found to extend the host range of Rhizobium leguminosarum bv. viciae to include nodulation of a pea mutant, cultivar Afghanistan; the nodX gene product specifies the addition of an acetyl group to the terminal N-acetylglucosamine residue at the reducing end of the pentasaccharide core of this signal molecule. Several other classes of bacterial glycoconjugate have also been shown by genetic analysis to be essential for normal nodule development and function: these include a capsular extracellular polysaccharide; lipopolysaccharide in the outer membrane; and cyclic glucans present in the periplasmic space. Potential functions for these glycoconjugates are discussed in the context of tissue and cell invasion by Rhizobium. Some plant components involved in symbiotic interactions have been identified by the analysis of nodule-specific gene expression (early nodulins). Several of the cDNA clones encoding these early nodulins specify proline-rich proteins that presumably correspond to cell wall glycoproteins or membrane arabinogalactan proteins. Other plant glycoconjugates have been identified using monoclonal antibodies as probes. A plant glycoprotein present in intercellular spaces has been identified as a component of the luminal matrix of infection threads. Because it attaches to the surface of bacteria and is itself susceptible to oxidative cross-linking, this glycoprotein may be involved in limiting the progress of microbial infections. Endocytosis of bacteria into the plant cytoplasm is apparently driven by direct interactions between the bacterial surface and the plasma membrane that is exposed within an unwalled infection droplet; glycoprotein and glycolipid components of the plant membrane glycocalyx have been defined using monoclonal antibodies. Differentiation of endosymbiotic bacteroids is preceded by differentiation of the plant-derived peribacteroid membrane which encloses the symbiosome compartment. Using a monoclonal antibody that identifies a group of plant membrane-associated, inositol-containing glycolipids, we have identified a very early marker for the differentiation of peribacteroid membrane from plasma membrane.

Meat quality and muscle fibre type characteristics of Southdown Rams from high and low backfat selection lines.

Characteristics of the meat of 15-18-month Southdown rams from lines selected for high or low backfat depths (assessed ultrasonically at position C over the last rib) were compared. Half of the carcasses were electrically stimulated (ES) and within each carcass post-mortem treatments chosen to produce effects on meat tenderness were ageing periods of 1 or 15 days (Semimembranosus), early or delayed chilling (Biceps femoris), and trimming of the s.c. fat cover (Longissimus dorsi). These treatments had the expected effects on shear values, but the sizes of the effects were little affected by selection line or ES treatment. Selection line did not have any direct effects on shear values, reflectance values at several wavelengths, waterholding capacity, cooking loss or sarcomere length. The Semitendinosus muscle had a higher proportion of predominantly oxidative fibres for the high-backfat line, based on succinic dehydrogenase activity (P < 0·05), but there was no line difference in alkaline-stable ATPase activity in the same muscle. Muscle fibre diameter was similar for the two lines.

Composition of the cell walls of Nicotiana alata Link et Otto pollen tubes.

Cell walls isolated from pollen of Nicotiana alata germinated in vitro contain glucose and arabinose as the predominant monosaccharides. Methylation analysis and cytochemical studies are consistent with the major polysaccharides being a (1→3)-ß-D-glucan (callose) and an arabinan together with small amounts of cellulose. The cell walls contain 2.8% uronic acids. Alcian blue stains the pollen-tube walls intensely at the tip, indicating that acidic polysaccharides are concentrated in the tip. Synthetic aniline-blue fluorochrome is specific primarily for (1→3)-ß-D-glucans and stains the pollen-tube walls, except at the tip. Protein (1.5%), containing hydroxyproline (2.4%), is present in the cell wall.