Theoretical Basis of Protocols for Seed Storage III. Optimum Moisture Contents for Pea Seeds Stored at Different Temperatures

In previous work, we demonstrated that there was an optimummoisture level for seed storage at a given temperature (Vertucciand Roos, 1990), and suggested, using thermodynamic considerations,that the optimum moisture content increased as the storage temperaturedecreased (Vertucci and Roos, 1993b). In this paper, we presentdata from a two year study of aging rates in pea (Pisum sativum)seeds supporting the hypothesis that the optimum moisture contentfor storage varies with temperature. Seed viability and vigourwere monitored during storage under dark or lighted conditionsat relative humidities between 1 and 90%, and temperatures between-5 and 65°C. The optimum moisture content varied from 0·015g H₂O g^-1 d.wt at 65°C to 0·101 g H₂O g^-1 d.wt at15°C under dark conditions and from 0·057 at 35°Cto 0·092 g H₂O g^-1 d.wt at -5°C under lighted conditions.Our results suggest that optimum moisture contents cannot beconsidered independently of temperature. This conclusion hasimportant implications for 'ultra-dry' and cryopreservationtechnologies.Copyright 1994, 1999 Academic Press Seed storage, seed aging, seed longevity, water content, temperature, glass, desiccation damage, ultradry, Pisum sativum L., pea, cryopreservation     

Cloning of Multiple Forms of Goldfish Vimentin: Differential Expression in CNS

Abstract: In efforts to determine the primary structure of intermediate filament proteins in the goldfish visual pathway, we isolated clones from a retinal λgt11 cDNA expression library that represent goldfish vimentin. We show that there are at least two forms of goldfish vimentin, designated as vimentin α and vimentin β. RNase protection assays indicate that vimentin α mRNA is expressed in low amounts in retina, optic nerve, and brain and in higher amounts in spinal cord. In contrast, vimentin β mRNA is expressed in low amounts in retina, optic nerve, brain, and spinal cord and in very high amounts in eye lens. Immunohistochemical studies show that in the optic nerve, vimentin α is mainly restricted to blood vessels, meninges, and septa. Light staining is observed with this antibody in an astrocytic glial pattern throughout the optic nerve. Two-dimensional gel analysis shows that all of these goldfish vimentins are low abundant components of optic nerve cytoskeletal preparations.     

Cloning and Expression of a 5-Hydroxytryptamine7 Receptor Positively Coupled to Adenylyl Cyclase

Abstract: In a number of different cell types, phosphorylation of a 63-kDa protein has been shown to increase rapidly in response to stimuli that lead to an increase in intracellular calcium. Here, a stimulus-sensitive protein at this molecular weight is identified in PC12 cells and rat cortical synaptosomes as phosphoglucomutase. In addition, the added phosphate is shown to be in an oligosaccharide terminating in phosphodiester-linked glucose. In synaptosomes, incorporated radioactivity, following incubation with [¹⁴C]glucose or the [β-³⁵S]phosphorothioate analogue of UDP-glucose, was found to increase within 5 s of stimulation and return to baseline within 25 s. Despite the many pathways utilizing glucose, this was the only detectable protein glycosylation observed in synaptosomes. These results indicate that cytoplasmic glycosylation is reversible and rapidly regulated, and suggest that phosphoglucomutase undergoes an alteration in function and/or topography in response to increases in intracellular calcium.     

Mutations in somePolycomb group genes ofDrosophila interfere with regulation of segmentation genes

Mutations in severalPolycomb (Pc) group genes cause maternal-effect or zygotic segmentation defects, suggesting thatPc group genes may regulate the segmentation genes ofDrosophila. We show that individuals doubly heterozygous for mutations inpolyhomeotic and six otherPc group genes show gap, pair rule, and segment polarity segmentation defects. We examined double heterozygous combinations ofPc group and segmentation mutations for enhancement of adult and embryonic segmentation defects.Posterior sex combs andpolyhomeotic interact withKrüppel ² and enhance embryonic phenotypes ofhunchback andknirps, andpolyhomeotic enhanceseven-skipped. Surprisingly, flies carrying duplications ofextra sex combs (esc), that were heterozygous for mutations ofeven-skipped (eve), were extremely subvital. Embryos and surviving adults of this genotype showed strong segmentation defects in even-numbered segments. Antibody studies confirm that expression ofeve is suppressed by duplications ofesc. However,esc duplications have no effect on other gap or pair rule genes tested. To our knowledge, this is only the second triplo-abnormal phenotype associated withPc group genes. Duplications of nine otherPc group genes have no detectable effect oneve. Expression ofengrailed (en) was abnormal in the central nervous systems of mostPc group mutants. These results support a role forPc genes in regulation of some segmentation genes, and suggest thatesc may act differently from otherPc group genes.     

Genetic analysis of the minimal replicon of the Lactococcus lactis subsp. lactis biovar diacetylactis citrate plasmid

Using a combination of mutagenesis with the transposon and polymerase chain reaction subcloning, the essential elements of the replication region of the Lactococcus lactis subsp. lactis biovar diacetylactis citrate plasmid have been identified. An open reading frame, coding for a protein with homology to Rep proteins from other Lactococcus plasmids, is essential. This protein is trans-acting and could not be replaced by the Rep protein from another Lactococcus plasmid. A second open reading frame immediately downstream from the first could be removed or inactivated with no apparent effect on plasmid replication. A region containing two 10 by direct repeats and three tandem repeats of a 22 by sequence, immediately upstream of the essential open reading frame, is also essential and probably includes the origin of replication. A 181-bp DNA fragment containing this region was sufficient to allow replication in Lactococcus if the trans-acting protein was provided on another replicon. Single-stranded replication intermediates could not be detected, suggesting that the citrate plasmid uses theta replication rather than rolling-circle replication.     

Structural and functional diversity among bacterial interspersed mosaic elements (BIMEs)

Palindromic units (PU or REP) were defined as 40-nucleotide DNA sequences which are highly repeated in the genome of several members of the Enterobacteriaceae. They were shown to be a constituent of the bacterial interspersed mosaic element (BIME), in which they are associated with other repetitive sequences. We report here that Escherichia coli PU sequences contain three motifs (Y, Z¹ and Z²), leading to the definition of two BIME families. The BIME-1 family, highly conserved over 145 nucleotides, contains two PUs (motifs Y and Z¹). The BIME-2 family contains a variable number of PUs (motifs Y and Z²). We present evidence, using band shift experiments, that each PU motif binds DNA gyrase with a different affinity. This suggests that the two families are functionally distinct.     

Carbohydrate receptor-mediated gene transfer to human T leukaemic cells

The mucin-type carbohydrate Tn cryptantigen (GalNAc1-O-Ser/Thr,where GalNAc is N-acetyl-D-galactosamine) is expressed in manycarcinomas, in haemopoietic disorders including the Tn syndrome,and on human immunodeficiency virus (HIV) coat glycoproteins,but is not expressed on normal, differentiated cells becauseof the expression of a Tn-processing galactosyltransferase.Using Jurkat T leukaemic cells which express high levels ofTn antigen due to deficient Tn galactosylation, we have establishedthe Tn antigen-mediated gene transfer and demonstrate the considerableefficiency of this approach. We used poly(L-lysine) conjugatesof the monoclonal antibody 1E3 directed against the Tn antigento deliver the luciferase and ß-galactosidase reportergenes to Jurkat cells by receptor-mediated endocytosis. Additionof unconjugated 1E3 reduced transfection efficiency in a concentration-dependentmanner and incubation with free GalNAc abolished DNA transfercompletely, indicating that gene delivery is indeed mediatedby the Tn antigen. Pre-treatment of Jurkat cells with Vibriocholerae sialidase, which uncovers additional Tn antigens, resultedin an improvement of gene transfection. Both human and chickenadenovirus particles attached to the DNA/polylysine complexstrongly augmented transgene expression. When the ß-galactosidase(lacZ) gene was delivered to Jurkat cells by Tn-mediated endocytosis,up to 60% of the cells were positive in the cytochemical stainusing 5-bromo-4-chloro-3-indolyl-ß-D-galactopyranoside(X-gal) as a chromogenic substrate. The efficiency of the transferrinreceptor-mediated DNA uptake into Jurkat cells was comparativelylow, although these cells were shown to express considerableamounts of transferrin receptor. We show here that a mucin-typecarbohydrate antigen mediates highly efficient DNA uptake byendocytosis into Jurkat T cells. This method represents a 50-foldimprovement of Jurkat cell transfection efficiency over otherphysical gene transfer techniques. Specific gene delivery toprimary cancer cells exhibiting Tn epitopes may especially bedesirable in immunotherapy protocols. adenovirus endocytosis gene transfer T cell Tn antigen     

Fish species composition before and after construction of a main stem reservoir on the White River,Colorado

Synopsis The completion in the fall of 1984 of Taylor Draw Dam on the White River, Colorado, formed Kenney Reservoir — thus impounding the last significant free-flowing tributary in the Upper Colorado River Basin. Fishes were sampled above and below the dam axis prior to closure of the dam and in the reservoir and river downstream following impoundment. While immediate effects of the dam to the ichthyofauna included blockage of upstream migration to 80 km of documented range for endangered Colorado squawfish, the reservoir also proved to have profound delayed effects on the river's species composition. Pre-impoundment investigations in 1983–1984 showed strong domination by native species above, within, and below the reservoir basin. By 1989–1990, non-native species comprised roughly 90% of the fishes collected in the reservoir and 80% of the fishes collected in the river below the dam. Initially, fathead minnow, whose numbers quickly increased in the new reservoir, dominated all post-impoundment collections, but red shiner became the most abundant fish collected in the river below the dam by 1989–1990. While agency stocking programs for the reservoir sought to emphasize a sport fishery for salmonids, primarily rainbow trout, local enthusiasm for warmwater sport fishes resulted in illicit transfers of these species from nearby impoundments. Several species, formerly rare or unreported in the White River in Colorado, including white sucker, northern pike, green sunfish, bluegill, largemouth bass and black crappie, were present in the river following impoundment. Our investigation indicates smaller-scale, main-stem impoundments that do not radically alter hydrologic or thermal regimes can still have a profound influence on native ichthyofauna by facilitating establishment and proliferation of nonnative species.Cooperators are the U.S. Fish and Wildlife, the Colorado Division of Wildlife, and Colorado State University