MPDA: Microarray pooled DNA analyzer

Background  

Microarray-based pooled DNA experiments that combine the merits of DNA pooling and gene chip technology constitute a pivotal advance in biotechnology. This new technique uses pooled DNA, thereby reducing costs associated with the typing of DNA from numerous individuals. Moreover, use of an oligonucleotide gene chip reduces costs related to processing various DNA segments (e.g., primers, reagents). Thus, the technique provides an overall cost-effective solution for large-scale genomic/genetic research. However, few publicly shared tools are available to systematically analyze the rapidly accumulating volume of whole-genome pooled DNA data.     

Global transcriptional response of pig brain and lung to natural infection by Pseudorabies virus

Background  

Pseudorabies virus (PRV) is an alphaherpesviruses whose native host is pig. PRV infection mainly causes signs of central nervous system disorder in young pigs, and respiratory system diseases in the adult.     

Effects of hypoxia on muscle protein synthesis and anabolic signaling at rest and in response to acute resistance exercise

Chronic reductions in tissue O(2) tension (hypoxia) are associated with muscle atrophy and blunted hypertrophic responses to resistance exercise (RE) training. However, the effect of hypoxia on muscle protein synthesis (MPS) at rest and after RE is unknown. In a crossover study, seven healthy men (21.4 ± 0.7 yr) performed unilateral leg RE (6 × 8 repetitions at 70% 1-repetition maximum) under normoxic (20.9% inspired O(2)) and normobaric hypoxic (12% inspired O(2) for 3.5 h) postabsorptive conditions. Immediately after RE the rested leg was biopsied, and a primed continuous infusion of [1,2-(13)C(2)]leucine was maintained for 2.5 h before final biopsies from both legs to measure tracer incorporation and signaling responses (i.e., ribosomal S6 kinase 1). After 3.5 h of hypoxia, MPS was not different from normoxia in the rested leg (normoxia 0.033 ± 0.016 vs. hypoxia 0.043 ± 0.016%/h). MPS increased significantly from baseline 2.5 h after RE in normoxia (0.033 ± 0.016 vs. 0.104 ± 0.038%/h) but not hypoxia (0.043 ± 0.016 vs. 0.060 ± 0.063%/h). A significant linear relationship existed between MPS 2.5 h after RE in hypoxia and mean arterial blood O(2) saturation during hypoxia (r(2) = 0.49, P = 0.04). Phosphorylation of p70S6K(Thr389) remained unchanged in hypoxia at rest but increased after RE in both normoxia and hypoxia (2.6 ± 1.2-fold and 3.4 ± 1.1-fold, respectively). Concentrations of the hypoxia-responsive mTOR inhibitor regulated in development and DNA damage-1 were unaltered by hypoxia or RE. We conclude that normobaric hypoxia does not reduce MPS over 3.5 h at rest but blunts the increased MPS response to acute RE to a degree dependent on extant SpO(2).     

The evolution of the reproductive system of Urodasys (Gastrotricha: Macrodasyida)

Macrodasyidan gastrotrichs are hermaphrodites with complex reproductive organs that function in sperm transfer and receipt, but homology among the organs of members of different clades remains undetermined, as does a broader understanding of evolutionary trends in the reproductive biology of macrodasyidans. In this study, we investigate the evolution of reproduction in Urodasys, a clade of 15 macrodasyidan species that shows variability in reproductive mode (hermaphroditic and parthenogenetic) and sexual anatomy. We use partial 18S rDNA sequence data from 30 specimens representing five described species, sequence data from one undescribed species in GenBank, and sequence data from a potentially new species found at Capron Shoal, Florida, to gain insight into the phylogeny of the clade and clarify evolutionary trends in reproductive modality. Based on a total of 33 specimens of seven potential species, we found that members of Urodasys can be separated into three clades reflective of different reproductive modalities: Clade I, species with paired male and female gonads but without accessory sexual organs; Clade II, species with a single left testis, paired ova, and accessory organs including a sclerotic stylet; and Clade III, parthenogenetic species without testes or accessory organs. In addition, we find that the potentially new species from Florida can form spermatophores, a condition shared with another species in Clade I. Herein, we describe this novel spermatophore‐bearing species and discuss the significance of spermatophore formation in the genus.     

Products of lipopolysaccharide-activated macrophages (tumor necrosis factor-alpha, transforming growth factor-beta) but not lipopolysaccharide modify DNA synthesis by rat trophoblast cells exhibiting the 80-kDa lipopolysaccharide-binding protein

Pregnancy losses from gram negative bacterial infections could be caused by direct effects of LPS on placental cells, or indirectly via LPS activation of macrophages in the uteroplacental unit. To evaluate those alternatives, LPS, LPS-activated peritoneal cells, conditioned medium from LPS-activated peritoneal cells, and some purified and recombinant molecules known to be secreted by activated macrophages were tested for their abilities to modify DNA synthesis by rat trophoblast cells. Three trophoblast cell lines derived from midgestation placentas of outbred and inbred rats were used for the experiments. Although the 80-kDa LPS-binding protein was demonstrated on trophoblast cells, LPS alone had no effect on the ability of trophoblast cells to synthesize DNA. In cocultures, trophoblast cell DNA synthesis was slightly enhanced by low concentrations of both unstimulated and LPS-activated peritoneal cells. At higher concentrations, LPS-activated cells caused significant inhibition of DNA synthesis by trophoblast cells. Conditioned media from LPS-activated peritoneal cells were highly inhibitory to trophoblast cell DNA synthesis. When specific molecules likely to be components of those media were tested, IL-1 was found to have a modest but reproducible stimulatory effect and PGE2 did not change trophoblast cell incorporation of [3H]TdR. In contrast, trophoblast cell DNA synthesis was markedly inhibited in a dose-dependent manner by both TNF-alpha and TGF-beta 1. No differences in the sensitivity of trophoblast cells from outbred and inbred rats were observed. Given the limitations of the experimental model system, the results suggest that in cases of infection by gram-negative bacteria LPS may have an adverse effect on pregnancy by stimulating resident macrophages to generate and release molecules that are inhibitory to trophoblast cell DNA synthesis.     

Identification of an S-locus glycoprotein allele introgressed from B. napus ssp. rapifera to B. napus ssp. oleifera.

We have previously described a developmentally regulated mRNA in maize that accumulates in mature embryos and is involved in a variety of stress responses in the plant. The sequence of the encoded 16 kDa protein (MA16) predicts that it is an RNA-binding protein, since it possesses a ribonucleoprotein consensus sequence-type RNA-binding domain (CS-RBD). To assess the predicted RNA binding property of the protein and as a starting point to characterize its function we have used ribohomopolymer-binding assays. Here we show that the MA16-encoded protein binds preferentially to uridine- and guanosine-rich RNAs. In light of these results a likely role for this protein in RNA metabolism during late embryogenesis and in the stress response is discussed.     

Isolation of a bacterium resembling Pirellula species from primary tissue culture of the giant tiger prawn (Penaeus monodon).

During attempts to establish tissue cultures from hepatopancreas, heart, and hemolymph of the giant tiger prawn (Penaeus monodon), using a medium including penicillin, streptomycin, and amphotericin B, bacterial contamination in the form of a sheet of growth attached to the tissue culture vessel was a persistent problem. Contaminant bacteria were teardrop-shaped cells arranged in rosettes, and electron microscopy revealed buds, crateriform structures, and the absence of a peptidoglycan layer in the cell wall, features characteristic of bacteria in the Planctomyces-Pirellula group, a phylogenetically distinct group of eubacteria. Two strains of contaminant bacteria were isolated in pure culture. Both exhibited morphology and antibiotic resistance consistent with their membership in the Planctomyces-Pirellula group (order Planctomycetales) of eubacteria. Tissue culture media for marine invertebrates may select for such bacteria if high concentrations of cell wall synthesis-inhibiting antibiotics are included.     

Molecular evolution of mitochondrial 12S RNA and cytochrome b sequences in the pantherine lineage of Felidae

DNA sequence comparisons of two mitochondrial DNA genes were used to infer phylogenetic relationships among 17 Felidae species, notably 15 in the previously described pantherine lineage. The polymerase chain reaction (PCR) was used to generate sequences of 358 base pairs of the mitochondrial 12S RNA gene and 289 base pairs of the cytochrome b protein coding gene. DNA sequences were compared within and between 17 felid and five nonfelid carnivore species. Evolutionary trees were constructed using phenetic, cladistic, and maximum likelihood algorithms. The combined results suggested several phylogenetic relationships including (1) the recognition of a recently evolved monophyletic genus Panthera consisting of Panthera leo, P. pardus, P. onca, P. uncia, P. tigris, and Neofelis nebulosa; (2) the recent common ancestry of Acinonyx jubatus, the African cheetah, and Puma concolor, the American puma; and (3) two golden cat species, Profelis temmincki and Profelis aurata, are not sister species, and the latter is strongly associated with Caracal caracal. These data add to the growing database of vertebrate mtDNA sequences and, given the relatively recent divergence among the felids represented here (1-10 Myr), allow 12S and cytochrome b sequence evolution to be addressed over a time scale different from those addressed in most work on vertebrate mtDNA.