Immunofluorescent and calcofluor white staining of developing tracheary elements inZinnia elegans L. Suspension cultures

Summary Xylogenesis has been studied in primary suspension cultures ofZinnia elegans L.: The wall patterns produced in culture closely resemble those described for intact tissues (annular, spiral, reticulate, scalariform, pitted). Using fluorescence microscopy and immuno-cytochemical techniques we have followed both the changes in wall deposition and microtubule organization during xylogenesis. Calcofluor white has been used to detect secondary wall deposition before it can be observed using either phase contrast or polarization optics. The development of tracheary elements can be divided into three stages: 1. microtubules grouped into bands without secondary wall deposition evident; 2. groups of microtubules subtending wall material only visible using Calcofluor white; 3. a complex microtubule pattern reflected by well developed wall thickenings detected using Calcofluor, phase contrast and polarization optics.     

Kinetics of Micronucleus Expression In Synchronized Irradiated Chinese Hamster Ovary Cells

The kinetics of expression of radiation-induced micronuclei (MN) in synchronized Chinese hamster cells (CHO) was examined. the purpose of the study was to determine if the cell cycle distribution of a population significantly influences the levels of radiation induced MN, thereby obscuring the exact quantification of the radiation effect. Cells were synchronized by centrifugal elutriation, irradiated, and then different phases of the cell cycle were examined for: cell cycle progression, division probability, and temporal expression of MN. the results demonstrate that the time interval for maximal MN expression is long enough that the position of cells in the cell cycle and radiation induced division delays do not prevent the majority of cells from completing their first post-irradiation mitosis, therefore, expressing MN. By following the progression of synchronized cell populations by flow cytometry and also examining the time of division of individual cells for 24 hr after irradiation, we observed that the maximum number of cells from all phases of the cell cycle are in their first post-irradiation interphase at that time, thus explaining the MN results.     

Comparative study of Agrobacterium biotypes 1, 2 and 3 by electrophoresis and serological methods

A comparative study by electrophoresis and serology of strains representing the three Agrobacterium biotypes was carried out. Thirteen Spanish isolates and strains from international collections were included. Ten antisera were prepared by using strains from the three biotypes and different types of antigens. The strains were studied by immunodiffusion, indirect immunofluorescence and indirect ELISA. Serological relationship among all the strains was observed, although serological heterogeneity within each of the biotypes occurred. Biotype 3 appears as serologically related to biotypes 1 and 2, having an intermediate position. This observation is in agreement with their biochemical characteristics. Electrophoretic analysis of the three biotypes showed that there was high variability. Three main bands appeared in the six strains studied. One specific band occurred in the biotype 1 strains and another in the biotype 3 strains.     

Giemsa stain applied to deplasticized sections to identify pancreatic islet cells

A new application of the Giemsa stain to demonstrate endocrine cells in deplasticized sections of Epon embedded material is described. Its application to the pancreas of Rana temporaria is illustrated. The technique does not require postfixation with OsO4 and is easily performed in 30 min. It allows the easy identification of three types of endocrine cells (A, B, and D). A cells, preferentially located at the islet periphery, stain purple-blue. B cells, which occupy the interior of the islet, display a lilac color. D cells give a strong purple color; they are located both in the periphery of the islets and scattered among acinar cells. Positive identification of the cell types was made by immunocytochemistry and electron microscopy.     

Effect of parasympathetic denervation on acetylcholine levels in the rat parotid gland. Is there an extraneuronal pool of acetylcholine?

Parasympathetic denervation of the rat parotid gland by avulsion of the auriculotemporal nerve caused a marked and lasting decrease in gland weight. Parasympathectomy did not change the levels of choline in the gland but decreased by 60% the levels of acetylcholine (ACh) ten days after surgery and 65% at 28 days. It is puzzling that relatively high levels of ACh remained after parasympathetic denervation. The presence of additional cholinergic nerves that innervate the gland, or pass through it en route to other structures may account for some of the remaining ACh. Also, Schwann cells from denervated nerves might have contributed to some of the ACh. The existence of an extraneuronal source of ACh is considered.     

Relationship of the replication and incompatibility genes of an IncFVI haemolysin plasmid with other F-like haemolysin plasmids

The replication and incompatibility region of the IncFVI plasmid pSU502 has been isolated by in vitro DNA manipulation as part of a 12.6 kb plasmid, denominated pSU503. Plasmid pSU503 was strongly incompatible with its parental plasmid, pSU1, but was fully compatible with the haemolytic plasmids pSU316 (IncFIII/IV), pHly152 (IncI2) and pSU233 (Inc-pSU233). Furthermore, the 6.9 kb EcoRI fragment of pSU503 which carries the replication and incompatibility determinants of pSU1 did not show any detectable homology (less than 70%) with any of the haemolysin-determining plasmids with which it is compatible. Thus, homologous haemolysin determinants have become linked to apparently unrelated replicons.     

Acyl carrier protein from Escherichia coli I. Aspects of the solution structure as evidenced by proton nuclear Overhauser experiments at 500 MHz

The downfield aromatic (6-8 ppm) and upfield ring current shifted methyl regions (1-0 ppm) in the proton nuclear magnetic resonance spectrum of acyl carrier protein (ACP) from Escherichia coli have been examined at 500 MHz by using nuclear Overhauser methods. The data are analyzed in terms of the secondary structural model of Rock & Cronan (1979) [Rock, C. O., & Cronan, J. E., Jr. (1979) J. Biol. Chem. 254, 9778-9785], which suggests the existence of four alpha-helical segments joined by three beta-turns, and a short coil at the C terminus of the protein. Nuclear Overhauser effects among Tyr-71, Ile-69, Ile-72, and His-75 allow refinement of the secondary structure of the C terminus. Nuclear Overhauser effects among Tyr-71, Phe-28, and three Ile's also place stringent limitations on the folding of the four alpha-helices. These data allow the proposal of a tertiary structural model for ACP.     

ENZYMES OF NUCLEIC ACID METABOLISM IN THE BRAINS OF YOUNG AND ADULT RATS

A number of precursors of RNA are incorporated several-fold more readily into the RNA of brain slices from 10-day-old rats than into RNA of slices from adult animals. The brains of the young animals show moderately higher levels of some of the anabolic enzymes of RNA metabolism including RNA polymerase (nucleosidetriphosphate: RNA nucleotidyltransferase; EC 2.7.7.6) and substantially lower levels of the degradative enzymes, the nucleoside phosphorylases. The data suggest that all the enzymes work in a concerted fashion to produce an increased rate of synthesis in young animals rather than that any single controlling enzymic event is responsible.