Optimal conditions for breaking medically important yeasts by an inexpensive and simple method

Conditions for breaking various medically important yeasts using glass beads, 30 ml Corex centrifuge tubes, and a Vortex mixer were determined. From 75–95% ofCandida hyphal cells and all species of yeasts exceptSporothrix schenckii were broken when 10 g of 0.45–0.50 mm glass beads, 50–300 mg of wet cells in 5 ml of buffer, and 90 s of vortexing were employed. Yeasts ofSporothrix schenckii broke more efficiently when 0.25–0.30 mm beads were used.     

Luteal phase defects: a possible relationship between short hyperthermic phase and sporadic sexual behavior in women

Sporadic (less than regular weekly) coital activity is associated with a particular type of infertility—the short hyperthermic (luteal) phase. This preliminary report is the first suggestion that a reduced frequency of human sexual activity might, itself, be associated with subfertile reproductive function.     

Differentiation of the myoepithelial cells of the rat submandibular gland in vivo and in vitro: an ultrastructural study

Cytodifferentiation of the myoepithelial cells (MEC) of the rat submandibular gland (SMG) was observed by studying the prenatal and postnatal development of the gland in vivo and in vitro by light and electron microscopy. The anlage of the SMG first appeared on the fourteenth day of gestation and, from its earliest inception, was surrounded by an intact basal lamina. Presumptive myoepithelial cells were first seen at 18 days of gestation coinciding with the onset of secretion in the rudiment. These cells were flattened, peripherally located and subjacent to the epithelial basal lamina. Initial deposition of cytofilaments in the MEC's was observed during the first three days following birth and fully matured cells were seen as early as one week after birth. Presumptive and immature MEC's were observed undergoing mitosis, but once cytofilament deposition had begun in the cells they did not divide. Myoepithelium developed in relation to embryonic secretory structures and were only observed surounding acini and intercalated ducts in the adult gland. New myoepithelial cells were formed as long as new acinar-intercalated duct units were formed. Myoepithelial cells did not produce secretory type granules at any time during their development or in their mature state. Development of the MEC's in vitro paralleled that in vivo and supported the above observations.     

Microtubules in maize leaf protoplasts in relation to donor tissue and in vitro culture

Summary Maize (Zea mays) leaf protoplasts were isolated from various leaves of two-week (4-leaf) seedlings and from sections of the third leaf blades. Microtubules (MTs) were visualized using immunofluorescence microscopy. Only freshly isolated protoplasts from the third and fourth leaf blades contained MTs, with protoplasts from the fourth leaf containing the most i.e. 13% of fourth-leaf protoplasts contained MTs. In general, protoplasts with fewer and smaller chloroplasts had more MTs. Initially 90–95% of protoplasts from basal portions of leaves had MTs but the percentage decreased slightly during culture particularly after 10 days. The antioxidant n-propyl gallate was beneficial in maintaining MT content. Few protoplasts from older sections intitially contained MTs but in all sections at least some protoplasts regained a significant MT content during culture (e.g., 10% of protoplast from the tip section possessed microtubules after 7 days of culture). Far fewer MTs were observed in individual leaf protoplasts than those isolated from suspension culture.Abbreviations BMS Black Mexican Sweet - MT microtubule - MtSB microtubule stabilizing buffer - PBS phosphate buffered saline     

DNA replication in maize leaf protoplasts

Maize leaf protoplasts were investigated for their metabolic competence and capacity to synthesize DNA. When protoplasts were incubated at elevated temperatures, they exhibited a heat shock response with specific proteins being preferentially synthesized. This indicated that the protoplasts were fully metabolically functional and capable of responding to environmental stimuli. Significant DNA synthesis was observed in these protoplasts after incorporation of ³H-thymidine into chromatin by trichloroacetic acid precipitation and by incorporation of 5-bromo-2-deoxyuridine (BrdU), an analog of thymidine, detected by immunofluorescence. The immunocytochemical method revealed that about 50% of nuclei in the maize leaf protoplasts were labelled after 3 days of culture and that most of these nuclei were labelled as intensely as normal mitotic cells. Aphidicolin, an inhibitor of DNA polymerase-, decreased the percentage of labelled nuclei, demonstrating that the labelling was substantially due to replicative DNA synthesis. However, chromosome condensation was not observed. It is proposed that these protoplasts are capable of DNA synthesis, but incapable of nuclear division. Effects of media additives on the number of nuclei entering S phase in these protoplasts were also assessed by the immunocytochemical method. Inclusion of 80mM Ca²⁺ in the enzyme solution increased protoplast yield and also appeared beneficial to DNA synthesis. The antioxidant, n-propyl gallate, which was used to stabilize the protoplasts, delayed the onset of DNA synthesis. Arginine and spermidine produced a slight increase in DNA synthesis.Abbreviations BrdU 5-bromo-2-deoxyuridine - DMSO dimethyl sulfoxide - n-PG n-propyl gallate - PBS phosphate-buffered saline Dedicated to Dr. Friedrich Constabel on the occasion of his 60th birthday     

Isolation of rsp-1, a novel cDNA capable of suppressing v-Ras transformation.

Using an expression cloning assay, we have isolated a novel cDNA, referred to as rsp-1, which suppresses the v-Ras-transformed phenotype. When introduced into NIH 3T3 fibroblasts under the control of a metallothionein promoter, rsp-1 confers resistance to v-Ras, but not to v-Mos or v-Src, and inhibits growth of the cells. The rsp-1 cDNA contains a 831-bp open reading frame encoding a 277-amino-acid leucine-rich protein. The rsp-1 cDNA exhibits no significant homology to sequences in the DNA data bases. However, searches of the protein data bases revealed that it contains a series of leucine-based repeats which are homologous to the leucine repeats found in the regulatory region of the yeast adenylyl cyclase. rsp-1 specific RNA is detectable in a wide variety of cell lines and tissues, and the gene is conserved among eukaryotic species. These data suggest that rsp-1 plays a role in Ras signal transduction.     

FACTORS AFFECTING RED SPRUCE REGENERATION IN DECLINING AREAS OF CAMELS HUMP MOUNTAIN,VERMONT

Scarcity of red spruce (Picea rubens Sarg.) seedlings in declining spruce-fir forests of Camels Hump mountain, Vermont, prompted a study on some contributing factors involved in failure of spruce regeneration. Cones were shorter than those from low elevation red spruce trees from unaffected sites. Seed number in cones collected in declining areas of Camels Hump was low as were seed sizes and weights. Seed germination was at control levels only in good seed years. Capacity of seeds to form seedlings was reduced relative to that of controls, although seedling growth was normal. Coniferous litter contains presumed allelopathic substances leachable by contemporary precipitations that affect seed germination and seedling root development in red spruce, but not in balsam fir. Shield fern contains leachable substances that reduce seed germination and seedling root development in red spruce, but not balsam fir. Roots of red spruce germlings have lower capacity to penetrate through the increased forest duff depths of declining forests than do balsam fir roots. It is anticipated that substantial reproduction of red spruce will not occur in declining montane conifer forests under present conditions.     

Proposal for adoption of the base of the Undulograptus austrodentatus Biozone as a global Ordovician stage and series boundary level

The base of the Undulograptus austrodentatus Biozone appears to be a synchronous event that is widely recognizable within graptolitic facies around the world. It occurs within an interval in which graptolite species ranges are now well known and in which there is a rapid turnover in the composition of graptolite faunas. This turnover reflects the rapid evolutionary radiation of the Diplograptacea simultaneously with the appearance of several distinctive pseudisograptid and glossograptid species. These events provide the basis for the recognition of two thin but widely applicable subzones; a lower Arienigraptus zhejiangensis Subzone and an upper U. sinicus Subzone. The occurrence of the lower boundary of the U. austrodentatus Biozone within a succession of first appearances also permits accurate and reliable identification of the boundary as well as assessment of stratigraphic completeness across the boundary interval in correlated sections. Diverse graptolite faunas of late Yapeenian and early Darriwilian age occur in association with the Histiodella altifrons Biozone of the North American midcontinent conodont zonation and the Paroistodus originalis and Microzarkodina parva biozones of the North Atlantic conodont zonation. They also occur in association with the shelly-fossil zonations developed for several different continents. These features of the base of the U. austrodentatus Biozone make it a suitable level for use as the boundary level for a global stage. Its stratigraphic position within the Ordovician System relative to other likely global stages as well as its coincidence with one of the major events in graptolite evolutionary history suggest that this level also may be a suitable level for the base of a global Middle Ordovician Series.Ordovician System, Ordovician stages, graptolite zonation, chronostratigraphy, international correlation. Charles E. Mitchell and Jörg Maletz, Department of Geology, State University of New York at Buffalo, Buffalo, New York 14260-1550, USA; 13th July, 1994; revised 22nd May, 1995.