Rhizogenesis in stem discs of Malus pumila rootstock M9 “Jork”: I. Hormonal and environmental effects on root induction and callus formation

To establish a protocol suited for the molecular characterization of root induction the influences of explant position, etiolation state and orientation as well as temperature and light intensity on root and callus formation were investigated. Depending on these factors stem discs of Malus incubated on indole-3-butyric acid containing medium and subsequently on hormone-free medium regenerated roots and callus of filamentous and smooth texture to a varying extent. Concentration and incubation duration of indole-3butyric acid strongly affected rooting and the production of smooth callus. Moreover smooth callus was profuse at the low light levels applied during rooting. Rooting efficiency decreased and filamentous callus increased between 19°C and 25°C. Explants close to the shoot apex displayed reduced rooting efficiency and profuse filamentous callus. There was a strong effect of explant orientation on root and filamentous callus formation.Abbrevations IBA indole-3-butyric acid - IAA indole-3-acetic acid - BA 6-benzylaminopurine - MS Murashigeand Skoog - rts roots - rt rooted - rtd rooted discs     

Evolution and ultrastructure of the bovine spermatogonia precursor cell line

We have used a cytochemical technique to investigate the distribution of acetylcholinesterase (AChE) activity in the deutocerebrum of the brain of the sphinx moth Manduca sexta. To distinguish between extra-and intracellular pools of the enzyme, some brains were treated prior to histochemical staining with echothiophate, an irreversible AChE inhibitor which penetrates cell membranes very slowly and, therefore, inhibits only extracellular AChE. In the antennal nerve, fascicles of presumably mechanosensory fibers show echothiophateinsensitive AChE activity. They bypass the antennal lobe and project to the antennal mechanosensory and motor center of the deutocerebrum. In the antennal lobe, fibers in the coarse neuropil, cell bodies in the lateral cell group, and all glomeruli exhibit AChE activity. In most ordinary glomeruli, echothiophate-sensitive AChE activity is concentrated in the outer cap regions, corresponding to the terminal arborizations of olfactory afferents. A previously unrecognized glomerulus in the ventro-median antennal lobe shows uniform and more intense AChE-specific staining that the other glomeruli. No AChE activity appeared to be associated with malespecific pheromone-sensitive afferents in the macro-glomerular complex. About 67 interneurons with somata in the lateral cell group of the antennal lobe show echo-thiophate-insensitive AChE activity. These neurous seem to be members of two types of antennal-lobe projection neurons with fibers passing through the outer-antenno-cerebral tract to the protocerebrum. AChE-stained arborizations of these neurons appear to invade all glomeruli, including three distinguishable subunits of the male-specific macroglomerular complex. In echothiophate-treated animals, the projections of one of these types of fiber form large terminals in the lateral horn of protocerebrum, which partly protrude into the adjacent glial cell layer. The results suggest that extracellularly accessible AChE is associated with ordinary olfactory receptor terminals but apparently not with pheromone-sensitive afferents. Intracellular AChE appears to be present in antennal mechanosensory fibers and in two types of olfactory projection neurons of the antennal lobe. The study provides further evidence for cholinergic neurotransmission of most antennal afferents. The AChE-containing interneurons might be cholinergic as well or use the enzyme for functions unrelated to hydrolysis of acetylcholine.Abbreviations ACh acetylcholine - AChE acetylcholinesterase - AL antennal lobe - AMMC antennal mechanosensory and motor center - ChAT choline acetyltransferase - IACT inner antenno-cerebral tract - MGC macroglomerular complex     

Beet curly top virus DI DNA-mediated resistance is linked to its size

Beet curly top virus (BCTV) infection is associated with the de novo synthesis of a heterogeneous population of subgenomic viral DNAs. Nicotiana benthamiana plants transformed with a partial repeat of one such subgenomic DNA remained susceptible to infection but produced ameliorated symptoms when agroinoculated with BCTV. Symptom amelioration is associated with the mobilization of subgenomic DNA from the integrated copy. In an attempt to improve the resistance, N. benthamiana has been transformed with a partial repeat of a much smaller subgenomic DNA. However, transgenic plants showed almost no resistance although subgenomic DNA was mobilised from the host genome. To further understand the molecular basis of the interference phenomenon, we compared the ability of BCTV to replicate and accumulate in leaf discs derived from resistant and non-resistant transgenic plants. Both subgenomic DNAs were able to interfere with virus replication but only in case of resistant plants the DI DNA efficiently suppressed viral accumulation.     

Fine Genetic Map of Mouse Chromosome 10 around the Polycystic Kidney Disease Gene,jcpk,and Ankyrin 3

A chlorambucil (CHL)-induced mutation of thejcpk(juvenile congenital polycystic kidney disease) gene causes a severe early onset polycystic kidney disease. In an intercross involvingMus musculus castaneus, jcpkwas precisely mapped 0.2 cM distal toD10Mit115and 0.8 cM proximal toD10Mit173.In addition, five genes,Cdc2a, Col6a1, Col6a2, Bcr,andAnk3were mapped in both thisjcpkintercross and a (BALB/c × CAST/Ei)F₁× BALB/c backcross. All five genes were eliminated as possible candidates forjcpkbased on the mapping data. Thejcpkintercross allowed the orientation of theAnk3gene relative to the centromere to be determined.D10Mit115, D10Mit173, D10Mit199,andD10Mit200were separated genetically in this cross. The order and genetic distances of all markers and gene loci mapped in thejcpkintercross were consistent with those derived from the BALB/c backcross, indicating that the CHL-induced lesion has not generated any gross chromosomal abnormalities detectable in these studies.     

System to study horizontal gene exchange among microorganisms without cultivation of recipients

Ribosomal RNA genes are characterized by highly conserved sequences and are present in multiple copies in most prokaryotic chromosomes. In principle, therefore, they might serve as sites for homologous recombination between unrelated microorganisms. Plasmids containing 23S ribosomal gene sequences, from different bacteria, which had been interrupted by insertion of a kanamycin-resistance gene, were used to transform Acinetobacter sp. DSM587 (former name: Acinetobacter calcoaceticus BD413-ivl10). In all cases, homologies between the 23S rRNA genes of phylogenetically distant bacteria and Acinetobac-ter sp. DSM587 were sufficient for replacement recombination events. The integration events, resulting in inactivation of any one of the seven rrn operons of Acinetobacter sp. DSM587, had no observable influence on cell growth. These results suggest the possibility of rRNA genes serving as natural vehicles for horizontal gene transfer. They also provide the basis of a novel strategy to analyse gene transfer without selection or cultivation of recipient cells. Because of the highly conserved structure of bacterial rrn operons, recombination events subsequent to gene transfer can be readily identified by polymerase chain reaction amplification of the recombinant sequence using a universal forward primer for the 16S rRNA gene and a reverse primer specific for the integrated marker gene.     

Morphology of the bovine Sertoli cell during the spermatogenetic cycle

Summary The morphology of the bovine Sertoli cell was studied during 6 different phases of the spermatogenetic cycle. Tubular dimensions do not vary significantly during the phases. Sertoli cells occupy 27.0% (phase 4) to 38.4% (phase 8) of the tubular epithelium. Sertoli cells of phase 1 are approximately 20% larger than during the other phases. 30–35% of Sertoli cell volume consists of organelles. Mitochondrial (about 5.0%) and nuclear (about 5.7%) volume densities remain remarkably stable during the cycle, irrespective of changes in Sertoli cell size. Phagocytic capacity of bovine Sertoli cells is only moderate. Elimination of excess spermatid cytoplasm occurs to a large extent prior to spermiation. The majority of spermatid residual bodies undergoes autolytic decay while attached to the Sertoli cell apical surface. Aggregates of densely packed cisternae of the smooth endoplasmic reticulum (ER) located in a basal position and associated with the acrosome-phase and maturation-phase spermatids contribute between 14 and 17% to Sertoli cell volume. During phase 3 the ER pinches off a large number of small, smooth-walled vesicles filled with flocculent content. The contact area between Sertoli cells and other tubular constituents changes considerably during the different phases. It is concluded that the blood-testis barrier is particularly impassable during phases 1 and 8. A lipid cycle does not exist in the bovine testicular tubular epithelium.     

THE CHEMICAL NATURE OF KERATOHYALIN GRANULES OF THE EPIDERMIS

Keratohyalin granules were isolated in the native form from the epidermis of newborn rats by the use of citric acid and a detergent. The isolated granules revealed a fine granular substructure in the electron microscope similar to that seen in situ. Analyses of amino acids by automated column-chromatography showed that proline and cystine are present in large proportions whereas histidine is present in a small amount. Accordingly, it was concluded that keratohyalin represents a sulfur-rich amorphous precursor of the horny cell content, rather than a sulfur-poor side product of the keratinization process, or a unique histidine-rich protein as proposed by in situ histochemical and radioautographic studies.     

Mu-induced rifamycin-resistant mutations not located in the rpoB gene of Escherichia coli

Summary A new class of rifamicin-resistant mutants of Escherichia coli was obtained by lysogenic insertions of bacteriophage Mu Amp DNA. Rifamycin resistance is closely linked to the ampicillin resistance conferred by the prophage. Mapping by conjugation with auxotrophic markers revealed that the rifamycin-resistant mutations are located between 28 and 37 min on the E. coli chromosome standard map, some distance from the rpoB gene at 89.5 min. The DNA-dependent RNA polymerase of these mutants is highly sensitive to rifampicin.     

Segmental organisation of the head in the embryo of Drosophila melanogaster

Summary Embryos of Drosophila melanogaster were irradiated in the presumptive head region with a UV-laser microbeam of 20 m diameter at two developmental stages, the cellular blastoderm and the extended germ band. The ensuing defects were scored in the cuticle pattern of the head of the first-instar larva, which is described in detail in this paper. The defects caused by irradiating germ band embryos when morphologically recognisable lobes appear in the head region were used to establish the segmental origin of various head structures. This information enabled us to translate the spatial distribution of blastoderm defects into a fate map of segment anlagen. The gnathal segments derive from a region of the blastoderm between 60% and 70% egg length (EL) dorsally and 60% and 80% ventrally. The area anterior to the mandibular anlage and posterior to the stomodaeum is occupied by the small anlagen of the intercalary and antennal segments ventrally and dorsally, respectively. The labrum, which originates from a paired anlage dorsally at 90% EL, is separated from the remaining head segments by an area for which we did not observe cuticle defects following blastoderm irradiation, presumably because those cells give rise to the brain. The dorsal and lateral parts of the cephalo-pharyngeal skeleton appear to be the only cuticle derivatives of the non-segmental acron. These structures derive from a dorso-lateral area just behind the putative brain anlage and may overlap the latter. In addition to the segment anlagen, the regions of the presumptive dorsal pouch, anterior lobe and post-oral epithelium, whose morphogenetic movements during head involution result in the characteristic acephalic appearance of the larva, have been projected onto the blastoderm fate map. The results suggest that initially the head of the Drosophila embryo does not differ substantially from the generalised insect head as judged by comparison of fate map and segmental organisation.