Diversity of uncultured microorganisms associated with the seagrass Halophila stipulacea estimated by restriction fragment length polymorphism analysis of PCR-amplified 16S rRNA genes.

The diversity of microorganisms associated with the leaves of the seagrass Halophila stipulacea in the northern Gulf of Elat was examined by culture-independent analysis. Microorganisms were harvested by a sonication treatment for total-community genomic DNA isolation. Oligonucleotides complementary to conserved regions in the 16S rRNA-encoding DNA (rDNA) of bacteria were used for PCR amplification. The 16S rDNA PCR products were subcloned and further characterized by a restriction fragment length analysis termed ARDRA (amplified rDNA restriction analysis). These analyses were carried out after reamplifying the cloned fragments with two primers binding symmetrically to the plasmid immediately on both sides of the cloned insert. Computer-aided clustering was performed after separate restriction analysis with enzymes HinfI and HpaII. By this method, 103 cloned 16S rDNA fragments were clustered into a total of 58 different groups. Sequence analysis of clones with an identical ARDRA pattern confirmed that members of an ARDRA group were closely related to each other. The sequenced clones were found to be affiliated with a marine snow-associated plastid-like rRNA clone and with a marine Hyphomonas strain, respectively. The method applied in this study could be useful for the routine study of other microbial communities of interest.     

THE EVOLUTION OF ASYMMETRY IN SEXUAL ISOLATION: A MODEL AND A TEST CASE

We constructed a model for the evolution of sexual isolation by extending Lande's (1981) model of sexual selection. The model predicts that asymmetric sexual isolation is a transient phenomenon, characteristic of intermediate stages of divergence in sexually selected traits. Unlike the Kaneshiro (1976, 1980) proposal, our model does not depend upon drift and the loss of courtship elements to produce asymmetries in sexual isolation. According to our model, the direction of evolution cannot be predicted from asymmetry in sexual isolation. We tested some features of the model using data from an experimental study of sexual isolation in the salamander Desmognathus ochrophaeus. We tested for sexual isolation between 12 allopatric populations and found significant asymmetry in sexual isolation in about a quarter of the test cases. The highest degrees of asymmetry were associated with intermediate levels of divergence. A curvilinear relationship between isolation asymmetry and divergence was predicted by our model and was supported by statistical analysis of the salamander data.     

Zygotic Lethal Mutations with Maternal Effect Phenotypes in Drosophila Melanogaster. II. Loci on the Second and Third Chromosomes Identified by P-Element-Induced Mutations

Screens for zygotic lethal mutations that are associated with specific maternal effect lethal phenotypes have only been conducted for the X chromosome. To identify loci on the autosomes, which represent four-fifths of the Drosophila genome, we have used the autosomal ``FLP-DFS' technique to screen a collection of 496 P element-induced mutations established by the Berkeley Drosophila Genome Project. We have identified 64 new loci whose gene products are required for proper egg formation or normal embryonic development.     

Purification and characterization of 1-SST, the key enzyme initiating fructan biosynthesis in young chicory roots (Cichorium intybus)

A genuine 1-SST (sucrose:sucrose 1-fructosy] transferase, EC 2.4.1.99) was purified and characterized from young chicory roots ( Cichorium intybus L. var. foliosum cv. Flash) by a combination of ammonium sulfate precipitation, concanavalin A affinity chromatography, anion and cation exchange chromatography. This protocol produced a 63-fold purification and a specific activity of 4.75 U (mg protein)⁻¹. The mass of the enzyme was 69 kDa as estimated by gel filtration. On SDS-PAGE apparent molecular masses of 49 kDa (α-subunit) and 24 kDa (β-subunit) were found. Further specification was obtained by MALDI-TOF MS detecting molecular ions at m/z 40109 and 19 896. These two fragments were also found on a western blot using an SDS-boiled chicory root extract and chicken-raised polyclonal antibodies against the purified 1-SST, indicating that the enzyme is a heterodimer in vivo. The N-terminus of chicory root 1-SST α-subunit was shown to be highly homologous with the cDNA-derived amino acid sequences from barley 6-SFT and a number of β-fructosyl hydrolases (in-vertases and fructan hydrolases). However, chicory root 1-SST properties could be clearly differentiated from those of chicory root 1-FFT (EC 2.4.1.100), chicory root acid invertase (EC 3.2.1.26) and yeast invertase. The enzyme mainly produced 1-kes-tose and glucose from physiologically relevant sucrose concentrations, indicating that this 1-SST is the key enzyme initiating fructan biosynthesis in vivo. However, like chicory root 1-FFT and barley 6-SFT, the enzyme also showed some β-fructofuranosi-dase activity (fructosyl transfer to water) at very low sucrose concentrations. Although sucrose clearly is the best substrate for the enzyme, some transferase and β-fructofuranosidase activity were also detected using 1-kestose as the sole substrate.     

A wound-inducible gene fromSalix viminalis coding for a trypsin inhibitor

A gene designatedswin1.1 has been isolated by screening aSalix viminalis genomic library with a heterologous probe,win3 fromPopulus. The region sequenced included the entire coding sequence for a protein with 199 amino acids plus the promoter and terminator. At the 5 end of the coding region is a sequence that encodes a hydrophobic region of 25–30 amino acids, that could form a signal peptide. A putative TATAA box and polyadenylator sequence were identified. Introns were absent. The gene product showed similarities with serine protease inhibitors from the Kunitz family and especially withwin3 from wounded leaves ofPopulus. Southern blot analysis indicated thatswin1.1 is a member of a clustered gene family,swin1. An oligonucleotide corresponding to the putative hypervariable region to-wards the carboxyl end when used as a probe in Southern hybridization showed high specificity forswin1.1. Expression of theswin1.1 gene was enhanced in wounded leaves. Theswin1.1 coding region without the signal sequence was highly expressed inEscherichia coli and the protein showed inhibitory activity against trypsin but at most slight activity against the other proteases tested. A systemically induced protein, SVTI, with inhibitor activity against trypsin, was isolated fromSalix leaves by affinity chromatography on a column of trypsin-Sepharose 4B and N-terminal sequenced. It corresponded with the translatedswin1.1 gene at 16 of the 19 amino acid sites, suggesting that SVTI is encoded by another member of theswin1 gene family.     

Somatic embryogenesis in conifers—A case study of induction and development of somatic embryos in Picea abies

Vegetatively propagated material offers many advantages over seed material in forest tree breeding research and in reforestation programmes. Evidence is accumulating to suggest that using somatic embryos in forestry is a viable option. However, before somatic embryos can be used optimally in forestry, basic research aimed at increasing the number of responsive genotypes as well as the age of the primary explant is needed. This in turn requires the establishment of a basic understanding of the physiological and molecular processes that underlie the development of somatic embryos. The functions of genes and their developmental and tissue specific regulation are studied using transient and stable transformation techniques.The process of somatic embryogenesis can be divided into different steps: (1) initiation of somatic embryos from the primary explant, (2) proliferation of somatic embryos, (3) maturation of somatic embryos and (4) plant regeneration. Cortical cells in the primary explant are stimulated to go through repeated divisions so that dense nodules are formed from which somatic embryos differentiate. The first formed somatic embryos continue to proliferate and give rise to embryogenic cell lines. Embryogenic cell lines of Picea abies can be divided into two main groups A and B, based on morphology, growth pattern and secretion of proteins. Our results suggest that extracellular proteins play a crucial role in embryogenesis of Picea abies. Somatic embryos from group A can be stimulated to go through a maturation process when treated with abscisic acid. Mature somatic embryos can develop into plants.Abbreviations ABA abscisic acid - BA N⁶-benzyladenine - 2,4-D dichlorophenoxy acetic acid     

MEASURING THE EFFECTS OF PAIRING SUCCESS,EXTRA-PAIR COPULATIONS AND MATE QUALITY ON THE OPPORTUNITY FOR SEXUAL SELECTION

Sexual selection can act through variation in the number of social mates obtained, variation in mate quality, or variation in success at obtaining extra-pair fertilizations. Because within-pair fertilizations (WPF) and extra-pair fertilizations (EPF) are alternate routes of reproduction, they are additive, rather than multiplicative, components of fitness. We present a method for partitioning total variance in reproductive success (a measure of the opportunity for selection) when fitness components are both additive and multiplicative and use it to partition the variance into components that correspond to each mechanism of sexual selection. Computer simulations show that extra-pair fertilizations can either increase or decrease total variance, depending on the covariance between within-pair and extra-pair success. Simulations also suggest that for socially monogamous species, extra-pair fertilizations have a greater effect than variation in mate quality or pairing status on the opportunity for selection. Application of our model to data gathered for a population of red-winged blackbirds (Agelaius phoeniceus) indicates that most of the variance in male reproductive success was attributable to within-pair sources of variance. Nevertheless, extra-pair copulations increased the opportunity for selection because males varied both in the proportion of their social young that they sired and in the number of extra-pair mates that they obtained. Furthermore, large and positive covariances existed between the number of extra-pair mates a male obtained and both social pairing success and within-pair paternity, indicating that, in this population, males preferred as social mates also were preferred as extra-pair mates.     

Genomic reorganization and disrupted chromosomal synteny in the siamang (Hylobates syndactylus) revealed by fluorescence in situ hybridization

We employed in situ hybridization (“chromosome painting”) of chromosome-specific DNA libraries of all human chromosomes to establish homologies between the human and siamang karyotypes (Hylobates syndactylus, 2n = 50). Numerous intra- and interchromosomal rearrangements have led to a massive reorganization of the siamang karyotype. There have been a minimum of 33 translocations. The 24 siamang autosomes are composed of 60 recognizable segments that show DNA homology to regions of the 22 human autosomes. Only two autosomes have not been involved in translocations. The siamang presents a case, in a primate closely related to humans, in which chromosome morphology and synteny are highly disturbed in a manner similar to that encountered among rodents. © 1995 Wiley-Liss, Inc.     

Starvation- and stationary-phase-induced acid tolerance in Escherichia coli O157:H7.

Stationary phase and the starvation of log-phase cells increased the acid tolerance of Escherichia coli O157:H7 strains. Although the degree of acid tolerance varied, the survival of most O157:H7 strains exceeded that of other, related, pathogens in a synthetic gastric fluid.