Comparison of vitrification and slow cooling for umbilical tissues

The tissue cryopreservation maintains the cellular metabolism in a quiescence state and makes the conservation possible for an indefinite period of time. The choice of an appropriate cryopreservation protocol is essential for maintenance of cryopreserved tissue banks. This study evaluated 10 samples of umbilical cord, from which small fragments of tissue (Wharton’s jelly and cord lining membrane) were subjected to two protocols of cryopreservation: slow cooling and vitrification. The samples were frozen for a period of time ranging from 5 to 78 days. The efficiency of cryopreservation was evaluated by testing cell viability, histological analysis, cell culture, cytogenetic analysis and comparison with the results of the fresh samples. The results showed that the slow cooling protocol was more efficient than the vitrification for cryopreservation of umbilical cord tissue, because it has caused fewer changes in the structure of tissue (edema and degeneration of the epithelium) and, despite the significant decrease cell viability compared to fresh samples, the ability of cell proliferation in vitro was preserved in most samples. In conclusion, this study showed that it is possible to cryopreserve small fragments of tissue from the umbilical cord and, to obtain viable cells capable of proliferation in vitro after thawing, contributing to the creation of a frozen tissue bank.     

Overcoming Efficiency Limitations of SnS‐Based Solar Cells

Thin‐film solar cells are made by vapor deposition of Earth‐abundant materials: tin, zinc, oxygen and sulfur. These solar cells had previously achieved an efficiency of about 2%, less than 1/10 of their theoretical potential. Loss mechanisms are systematically investigated and mitigated in solar cells based on p‐type tin monosulfide, SnS, absorber layers combined with n‐type zinc oxysulfide, Zn(O,S) layers that selectively transmit electrons, but block holes. Recombination at grain boundaries is reduced by annealing the SnS films in H₂S to form larger grains with fewer grain boundaries. Recombination near the p‐SnS/n‐Zn(O,S) junction is reduced by inserting a few monolayers of SnO₂ between these layers. Recombination at the junction is also reduced by adjusting the conduction band offset by tuning the composition of the Zn(O,S), and by reducing its free electron concentration with nitrogen doping. The resulting cells have an efficiency over 4.4%, which is more than twice as large as the highest efficiency obtained previously by solar cells using SnS absorber layers.     

Inbreeding depression and selfing rate of Ipomoea hederacea var. integriuscula (Convolvulaceae)

Inbreeding depression and selfing rate were investigated in the self-compatible vine Ipomoea hederacea to assess the variability of the breeding system. Inbreeding depression differed between populations and the magnitude varied at germination, growth (as measured by aboveground biomass), and reproductive potential. Plants from Macon County, Alabama, USA, had significant inbreeding depression (31%) at germination, but no significant inbreeding depression for aboveground biomass or number of reproductive structures (buds and flowers) at 45 d post germination in the greenhouse or in the field. Plants from Morgan County, Alabama, however, had significant inbreeding depression (>50%) for all three stages in the greenhouse. In allozyme comparisons, five of the 11 I. hederacea populations surveyed had high selfing rates (66.66-92.53%) and high levels of homozygosity (F(IS) = 0.500-0.861) in 2003, and three of four populations surveyed in 2004 had selfing rates that exceeded 50%. High selfing rates, high levels of homozygosity, and low levels of inbreeding depression suggest that inbreeding depression may not present a significant barrier to the transmission of selfing alleles in some populations of I. hederacea, but does not account for the maintenance of a mixed mating system in other populations.     

Mapping nutrient resorption efficiencies of subarctic cryptogams and seed plants onto the Tree of Life

Nutrient resorption from senescing photosynthetic organs is a powerful mechanism for conserving nitrogen (N) and phosphorus (P) in infertile environments. Evolution has resulted in enhanced differentiation of conducting tissues to facilitate transport of photosynthate to other plant parts, ultimately leading to phloem. Such tissues may also serve to translocate N and P to other plant parts upon their senescence. Therefore, we hypothesize that nutrient resorption efficiency (RE, % of nutrient pool exported) should correspond with the degree of specialization of these conducting tissues across the autotrophic branches of the Tree of Life. To test this hypothesis, we had to compare members of different plant clades and lichens within a climatic region, to minimize confounding effects of climatic drivers on nutrient resorption. Thus, we compared RE among wide‐ranging basal clades from the principally N‐limited subarctic region, employing a novel method to correct for mass loss during senescence. Even with the limited numbers of species available for certain clades in this region, we found some consistent patterns. Mosses, lichens, and lycophytes generally showed low RE_N (<20%), liverworts and conifers intermediate (40%) and monilophytes, eudicots, and monocots high (>70%). RE_P appeared higher in eudicots and liverworts than in mosses. Within mosses, taxa with more efficient conductance also showed higher RE_N. The differences in RE_N among clades broadly matched the degree of specialization of conducting tissues. This novel mapping of a physiological process onto the Tree of Life broadly supports the idea that the evolution of conducting tissues toward specialized phloem has aided land plants to optimize their internal nitrogen recycling. The generality of evolutionary lines in conducting tissues and nutrient resorption efficiency needs to be tested across different floras in different climatic regions with different levels of N versus P availability.     

An evolutionary perspective on the resistance of Daphnia to the epizoic rotifer Brachionus rubens

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Callus formation and differentiation in tissue cultures of normal and cytoplasmic male sterile sorghum,pepper, sunflower,and tobacco

Summary The primary goal of this study was to establish callus cultures of selected plant taxa containing both normal (N) and cytoplasmic male-sterile (CMS) lines. A secondary goal was to attempt to differentiate the calli into whole flowering plants. Undifferentiated and organ differentiated calli were produced in specific lines of N and CMS sorghum, pepper, sunflower, and tobacco. Calli from both N and CMS lines of a given species developed similarly and grew well. Even though whole flowering plants have not yet been produced from any of the lines, this approach allows for good production of callus for studies dealing with CMS. Journal Paper No. J-7741 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa. Project No. 1914.     

Minor labdane diterpenes from Marrubium thessalum

Five new labdane diterpenes, thessalines A, B, and D, 14β-hydroxythessaline A, and 14β-hydroxythessaline B (1-5, resp.) were isolated from the aerial parts of Marrubium thessalum, along with the known labdane diterpene deacetylvitexilactone (6) and the methoxylated flavones 4',7-dimethylapigenin and salvigenin. (3S,5R)-Loliolide was also found in the same source. Their structures were established by 1D- and 2D-NMR (COSY, HSQC, HMBC, NOESY, and ROESY) and MS analyses. The plant produces a great variety of labdane-type diterpenes, with variations in functionalities, particularly in the side chain. Their structures could be of chemotaxonomic significance for the genus Marrubium.     

Recognition and separation of single particles with size variation by statistical analysis of their images

Macromolecules may occupy conformations with structural differences that cannot be resolved biochemically. The separation of mixed molecular populations is a pressing problem in single-particle analysis. Until recently, the task of distinguishing small structural variations was intractable, but developments in cryo-electron microscopy hardware and software now make it possible to address this problem. We have developed a general strategy for recognizing and separating structures of variable size from cryo-electron micrographs of single particles. The method uses a combination of statistical analysis and projection matching to multiple models. Identification of size variations by multivariate statistical analysis was used to do an initial separation of the data and generate starting models by angular reconstitution. Refinement was performed using alternate projection matching to models and angular reconstitution of the separated subsets. The approach has been successful at intermediate resolution, taking it within range of resolving secondary structure elements of proteins. Analysis of simulated and real data sets is used to illustrate the problems encountered and possible solutions. The strategy developed was used to resolve the structures of two forms of a small heat shock protein (Hsp26) that vary slightly in diameter and subunit packing.     

Development of a cellular assay system to study the genome replication of high- and low-risk mucosal and cutaneous human papillomaviruses

We found that recircularized high-risk (type 16 and 18) and low-risk mucosal (type 6b and 11) and cutaneous (type 5 and 8) human papillomavirus (HPV) genomes replicate readily when delivered into U2OS cells by electroporation. The replication efficiency is dependent on the amount of input HPV DNA and can be followed for more than 3 weeks in proliferating cell culture without selection. Cotransfection of recircularized HPV genomes with a linear G418 resistance marker plasmid has allowed subcloning of cell lines, which, in a majority of cases, carry multicopy episomal HPV DNA. Analysis of the HPV DNA status in these established cell lines showed that HPV genomes exist in these cells as stable extrachromosomal oligomers. When the cell lines were cultivated as confluent cultures, a 3- to 10-fold amplification of the HPV genomes per cell was induced. Two-dimensional (2D) agarose gel electrophoresis confirmed amplification of mono- and oligomeric HPV genomes in these confluent cell cultures. Amplification occurred as a result of the initiation of semiconservative two-dimensional replication from one active origin in the HPV oligomer. Our data suggest that the system described here might be a valuable, cost-effective, and efficient tool for use in HPV DNA replication studies, as well as for the design of cell-based assays to identify potential inhibitors of all stages of HPV genome replication.