Identification of a genetic contamination in a commercial mouse strain using two panels of polymorphic markers

Rapid detection of genetic contamination is critical in mouse studies involving inbred strains. During a Quantitative Trait Locus (QTL) study using simple sequence length polymorphism (SSLP) markers, we noticed heterozygosity at some loci of a commercially available inbred C57BL/6N mouse strain, suggesting a contamination by another mouse strain. A panel of 100 single-nucleotide polymorphism (SNP) markers was used to confirm and specify the genetic contamination suspected. Retrospective analyses demonstrated that the contamination took place as early as autumn 2003 and has persisted ever since at a fairly constant level. Contaminating alleles most probably originated from a DBA strain. Our data demonstrate the suitability of SNP markers for rapid detection and identification of the source of genetic contamination. Further, our results show the importance of a state-of-the-art genetic monitoring of the authenticity of murine inbred strains.     

Behaviour of granular starches in low-pressure glow plasma

Granular starches of nine botanical origins were exposed to low-pressure glow plasma generated in the air from either a high voltage DC supply or an AC inductor. Experiments with low energy air-plasma (10⁻² Torr) and starches provided insight into some details of their surface structure. Starches were partly oxidized to carboxylic starches and partly depolymerized. Their affinity to plasma depended on their botanical origin. The structure of starch granules seemed to be a crucial factor, which determined the behaviour of starches in plasma. Lipids and/or proteins stabilized polysaccharides to plasma being preferentially decomposed.     

Purification and characterization of D-glycerate-3-kinase from maize leaves

Glycerate kinase (GK; EC 2.7.1.31) from maize (Zea mays L.) leaves was purified by a sequence of ammonium-sulfate precipitations and chromatography on diethylaminoethyl-cellulose, hydroxyapatite, Sephadex G-75SF and dye ligand (Green A) columns. The purest preparation was almost 1300-fold enriched and had a specific activity of 68 mol · min^-1 · (mg protein) ^-1. The enzyme was a monomer of a relative molecular mass (M_r) of 44 kDa (kdalton) as determined by gel filtration, electrophoresis in dissociating conditions and by immunoblots. The enzyme was only weakly recognized by polyclonal antibodies against purified spinach GK, indicating substantial differences in molecular structure of the two proteins. Highly reducing conditions stabilized GK activity and were required for activation of crude leaf enzyme. The enzyme had a broad pH optimum of 6.8–8.5, and formed 3-phosphoglycerate and ADP as reaction products. Apparent K _ms for D-glycerate and Mg-ATP were 0.11 and 0.25 mM, respectively. The enzyme was strongly affected by a number of phosphoesters, especially by 3-phosphoglycerate (K _i= 0.36 mM), fructose bisphosphates and nucleoside bisphosphates. Inhibition by 3-phosphoglycerate was competitive to Mg-ATP and noncompetitive to D-glycerate. Pyruvate was found noncompetitive to D-glycerate (K _is=4 mM). The ratio of stromal concentration of Mg-ATP to phosphoesters, particularly to 3-phosphoglycerate, may be of importance in the regulation of GK during C₄-photosynthesis.Abbreviations DEAE diethylaminoethyl - kDa kdalton - GAP-DH glyceraldehyde phosphate dehydrogenase - GK glycerate kinase - LDH lactate dehydrogenase - 2-ME 2-mercaptoethanol - M_r relative molecular mass - PEP phosphoenolpyruvate - PGA(PK) phosphoglycerate (phosphokinase) - PK pyruvate kinase - SDS-PAGE sodium dodecyl sulfatepolyacrylamide gel electrophoresis     

Population growth and production of Habrotrocha rosa Donner (Rotifera: Bdelloidea) and its contribution to the nutrient supply of its host,the northern pitcher plant,Sarracenia purpurea L. (Sarraceniaceae)

The population growth and biomass production of the pitcher-plant (Sarracenia purpurea L.) inquiline, Habrotocha rosa Donner (Rotifera: Bdelloidea), its consumption by other pitcher-plant inqulines, and its excretion of phosphorus (PO4–P) and nitrogen (NO3–N and NH4–N), were investigated in laboratory experiments. Observed population growth and production rate of H. rosa were higher at pH 4 (2.3 rotifers d-1) than at pH 3 (1.3 rotifers d-1), 5 (1.9 rotifers d-1), or 6 (0.8 rotifers d-1). Populations of H. rosa are an abundant and reliable food source for larvae of the dipteran inqulines Wyeomyia smithii (Coq.) and Blaesoxipha fletcheri (Aldrich) that co-occur with H. rosa in S. purpurea pitchers. Abundance of H. rosa within a pitcher is negatively associated with abundance of dipteran larvae, and these larvae consume rotifers in direct proportion to rotifer density (Type I functional response). Habrotrocha rosa may also account for the majority of the plant's supply of N and P. An average population of rotifers in the field (∼400 per pitcher) can excrete ∼5.2 μg NO3-N, ∼3.91 μg NH4-N, and ∼18.4 μg PO4–P per day into a single leaf, and excretion rate is independent of water pH. Over the six-month growing season of pitcher-plants in Massachusetts, U.S.A., we estimate that rotifers could supply 8.8–43 mg of N and 18.2–88 mg of P. These values far exceed the amount of N and P previously estimated to be supplied annually to the plants through insect capture or rainfall. This revised version was published online in September 2006 with corrections to the Cover Date.     

Molecular characterization of multiple cDNA clones for ADP-glucose pyrophosphorylase from Arabidopsis thaliana

PCR amplification of cDNA prepared from poly(A)⁺ RNA from aerial parts of Arabidopsis thaliana, using degenerate nucleotide primers based on conserved regions between the large and small subunits of ADP-glucose pyrophosphorylase (AGP), yielded four different cDNAs of ca. 550 nucleotides each. Based on derived amino acid sequences, the identities between the clones varied from 49 to 69%. Sequence comparison to previously published cDNAs for AGP from various species and tissues has revealed that three of the amplified cDNAs (ApL1, ApL2 and ApL3) correspond to the large subunit of AGP, and one cDNA (ApS) encodes the small subunit of AGP. Both ApL1 and ApS were subsequently found to be present in a cDNA library made from Arabidopsis leaves. All four PCR products are encoded by single genes, as found by genomic Southern analysis.     

Stable co-transformation of maize protoplasts with gusA and neo genes

An efficient co-transformation protocol using polyethylene glycol was developed for Zea mays L. (cv. A188 × BMS) protoplasts isolated from suspension culture cells. Co-transformation was accomplished by using plasmid constructions containing -glucuronidase (gusA) or neomycin phosphotransferase (neo) gene coding sequences; both were under control of the CaMV 35S promoter. Protoplast culture and transformation conditions were optimized to assure efficient recovery of transformed cells. The overall efficiency of transformation was 1 × 10^–4 (calculated per viable protoplast plated). Among kanamycin-resistant lines, 50% showed a high level of GUS activity (above one unit). Southern blot hybridization confirmed the presence of numerous gusA and neo coding sequences in the maize genome. In two analyzed lines, integrated sequences appeared to be organized in tandem head-to-tail repeats. Results also indicated that the integrated sequences were partially methylated.