Aerobic Biodegradation of Crude Oil Components by Acidophilic Mycobacteria

Biodegradation of crude oil components by strain AG_S10, an acidophilic member of the genus Mycobacterium, was studied under extremely acidic conditions (pH 2.5). The degree of degradation of the same hydrocarbons in different kinds of oil was found to be different. The direction of biodegradation was, however, the same: the share of n-alkanes in oxidized oil decreased, while the share of branched alkanes increased. At the same time, the degree of redistribution of methane hydrocarbons in degraded oil varied significantly for different oils, although no strict dependence on the type of oil was found. After 28 days of incubation at 30°C and pH 2.5, the degradation of n- and iso-alkanes was 99 and 44%, respectively for the light, low-viscosity oil of the Nizhnevartovsk deposit, 58 and 32%, respectively for the medium-density oil of the Moscow oil-procesing plant, and 80 and 16% and 99 and 69%, respectively for the heavy, viscous oils of the Cheremukhovskoe and Usinkoye oil fields. Moreover, after extended cultivation time strain AG_S10 completely utilized alkanes, as well as a significant part of the naphthene component of the aliphatic fraction. The studied strain was characterized by ability to oxidize a broad spectrum of methane hydrocarbons, including high-molecular C₁₇–C₃₀n-alkanes, in oils of different properties and composition. Apart from its scientific interest, farther investigation of biodegradation of high-paraffin oils and viscous oils with elevated paraffin content by strain AG_S10 may be useful in view of the technical issues associated with paraffin accumulation in the course of recovery and transportation of these oils.     

Biodegradation of textile azo dye by <Emphasis Type="Italic">Shewanella decolorationis</Emphasis> S12 under microaerophilic conditions

The complete biodegradation of azo dye, Fast Acid Red GR, was observed under microaerophilic conditions by Shewanella decolorationis S12. Although the highest decolorizing rate was measured under anaerobic condition and the highest biomass was obtained under aerobic condition, a further biodegradation of decolorizing products can only be achieved under microaerophilic conditions. Under microaerophilic conditions, S. decolorationis S12 could use a range of carbon sources for azo dye decolorization, including lactate, formate, glucose and sucrose, with lactate being the optimal carbon source. Sulfonated aromatic amines were not detected during the biotransformation of Fast Acid Red GR, while H₂S formed. The decolorizing products, aniline, 1,4-diaminobenzene and 1-amino-2-naphthol, were followed by complete biodegradation through catechol and 4-aminobenzoic acid based on the analysis results of GC-MS and HPLC.     

DNAase I and cellular factors that affect chromatin structure

The use of DNAase I as a probe of chromatin structure is frequently fraught with problems of irreproducibility. We have recently evaluated this procedure, documented the sources of the problems, and standardized the method for reproducible results (Prentice and Gurley (1983) Biochim. Biophys. Acta 740, 134–144). We have now used this probe to detect differences in chromatin structure between cells blocked (1) in G₁ phase by isoleucine deprivation, or (2) in early S phase by sequential use of isoleucine deprivation followed by release into the presence of hydroxyurea. The cells blocked in G₁ phase have easily-digestible chromatin, while cells blocked in early S phase have chromatin which is much more resistant to DNAase I. These differences were found to be the result of diffusible factors found in the cytoplasm and nuclei of G₁- and S-phase cells, respectively. The G₁ cells contained a cytoplasmic factor which modulates the chromatin structure of S-phase nuclei to a more easily digestible state, while cells blocked in S phase contain a nuclear factor which modulates the chromatin structure of G₁ nuclei to a state more resistant to digestion. DNAase I is much more sensitive to these cell cycle-specific chromatin changes than is micrococcal nuclease. The results indicate that, under controlled conditions, DNAase I should be a valuable probe for detecting chromatin structural changes associated with cell cycle traverse, differentiation, development, hormone action and chemical toxicity.     

Flower-specific expression of <Emphasis Type="Italic">Arabidopsis</Emphasis><Emphasis Type="Italic">PCS1</Emphasis> driven by <Emphasis Type="Italic">AGAMOUS</Emphasis> second intron in tobacco decreases the fertility of transgenic plants

The PROMOTION OF CELL SURVIVAL 1 (PCS1) gene, encoding an aspartic protease, has an important role in determining the fate of cells in embryonic development and reproduction processes in Arabidopsis. To explore the potential function of the PCS1 gene in generating reproductive sterility, we placed the PCS1 gene under the control of an 1,869-bp nucleotide sequence from the 3′ end of the second intron (AG-I) of Arabidopsis AGAMOUS and CaMV 35S (–60) minimal promoter [AG-I-35S (–60)::PCS1], and introduced it into tobacco. RT–PCR results demonstrated that the PCS1 gene driven by AG-I-35S (–60) chimeric promoter was expressed only in anthers and carpels in the reproductive tissues of transgenic tobacco. Compared to wild-type plants, all AG-I-35S (–60) and AG-I-35S (–60)::PCS1 transgenic lines showed a normal phenotype throughout the vegetative growth phase. However, during the reproductive stage, most AG-I-35S (–60)::PCS1 transgenic plant anthers displayed delayed dehiscence, failed dehiscence, petalody and hypoplasia, and the pollen grains had different shapes and sizes with a distorted, shrunken, or collapsed morphology. Moreover, three transgenic lines, PCS1-1, PCS1-3 and PCS1-4, showed higher sterility than wild-type and AG-I-35S (–60) transgenic plants, respectively. These results showed that the construct of AG-I-35S (–60)::PCS1 was partially effective at preventing seed set and provided a novel sterility strategy.     

The afterglow thermoluminescence band as an indicator of changes in the photorespiratory metabolism of the model legume Lotus japonicus

The afterglow (AG) luminescence is a delayed chlorophyll fluorescence emitted by the photosystem II that seems to reflect the level of assimilatory potential (NADPH+ATP) in chloroplast. In this work, the thermoluminescence (TL) emissions corresponding to the AG band were investigated in plants of the WT and the Ljgln2‐2 photorespiratory mutant from Lotus japonicus grown under either photorespiratory (air) or non‐photorespiratory (high concentration of CO₂) conditions. TL glow curves obtained after two flashes induced the strongest overall TL emissions, which could be decomposed in two components: B band (t_max = 27–29°C) and AG band (t_max = 44–45°C). Under photorespiratory conditions, WT plants showed a ratio of 1.17 between the intensity of the AG and B bands (I_AG/I_B). This ratio increased considerably under non‐photorespiratory conditions (2.12). In contrast, mutant Ljgln2‐2 plants grown under both conditions showed a high I_AG/I_B ratio, similar to that of WT plants grown under non‐photorespiratory conditions. In addition, high temperature thermoluminescence (HTL) emissions associated to lipid peroxidation were also recorded. WT and Ljgln2‐2 mutant plants grown under photorespiratory conditions showed both a significant HTL band, which increased significantly under non‐photorespiratory conditions. The results of this work indicate that changes in the amplitude of I_AG/I_B ratio could be used as an in vivo indicator of alteration in the level of photorespiratory metabolism in L. japonicus chloroplasts. Moreover, the HTL results suggest that photorespiration plays some role in the protection of the chloroplast against lipid peroxidation.     

Genetic and physiological evidence concerning the development of chemically sensitive voltage-dependent ionophores in L6 cells

The electrophysiological properties of a tissue culture muscle line, L₆, and a K⁺ resistant mutant (MK₁) derived from L₆ were determined to elucidate certain aspects of membrane differentiation and function. MK₁ was selected as a clone of myoblasts resistant to the toxic effects of 55 mM K⁺. The resting potentials of L₆ and MK₁ myoblasts and myotubes were K⁺ dependent and equal. The amplitudes of the action potentials were equal in normal medium, but 27.7 mM K⁺ interfered with or eliminated the ability of L₆ myotubes to produce action potentials. MK₁ myotubes produced nearly normal action potentials under these conditions. Thus, the K⁺ resistant myoblasts differentiate into myotubes which have an action potential generating mechanism much less sensitive to K⁺ than the normal mechanism. Also, both d-tubocurarine and α-bungarotoxin enhance the amplitude of the action potentials produced by L₆ myotubes in the presence of 27.7 mM K⁺; these compounds do not enhance the amplitude of the action potentials produced by MK₁ myotubes under the same conditions. It is proposed that as a consequence of differentiation a type of ionophore present in myoblasts becomes a voltage-dependent ionophore in myotubes. Furthermore, these voltage-dependent ionophores can be chemically sensitive.     

Beta structure of periodic copolypeptides of L-alanine and glycine. Their relevance to the structure of silks

The β structure of seven periodic copolypeptides of l-alanine and glycine² namely: A₂G, AG, A₂G₂, A₃G₃, A₂G₃, AG₂ and AG₃ has been studied by infrared, X-ray and electron diffraction techniques. The sheets are made of anti-parallel chains and intersheet spacing is observed to depend both on residue sequence and composition. Samples with glycine molar fractions varying from 0.33 to 0.6 are found to be good models for group II of silk fibroins. The structure of Bombyx mori silk fibroin is discussed in the light of these new data.     

Biodegradation kinetics of trichloroethylene and1,2-dichloroethane by Burkholderia (Pseudomonas)cepacia PR131 and Xanthobacter autotrophicus GJ10.

The biodegradation kinetics for chlorinated aliphatic hydrocarbons trichloroethylene (TCE) by Burkholderia (Pseudomonas) cepacia PR1₃₁ and for1,2-dichloethane (1,2-DCA) by Xanthobacter autotrophicus GJ10 were determinedusing an initial rate method to determine the applicable rate law and relevant kinetic parametersunder aerobic conditions. A first order linear rate law applied to 1,2-DCA biodegradation by X. autotrophicus GJ10. The first order rate constant was determined to be 0.014 ml/min/mg.A non-linear rate law applied to TCE biodegradation by B. cepacia PR1₃₁.The maximum specific degradation rate constant was determined to be 0.8 nmol/min/mg protein,and the half saturation constant was determined to be 0.026 mM (3.47 ppm). Error analysisperformed on our analytical methods and computations, using a logarithmic differentiationmethod, indicated the relative error of our reported rate constants to be approximately 17%.Knowledge of the kinetic rate laws and kinetic parameters governing the biodegradation of TCEand 1,2-DCA by these strains will further the application of these strains in the environmental fieldand in waste treatment applications.     

Enhanced Biodegradation of Diesel Oil in Seawater Supplemented with Nutrients

The imbalance of C, N, and P caused by the spilled oil could be regulated by the addition of nitrogen and phosphorous. Moreover, different kinds of N and P sources were used in order to stimulate oil biodegradation under laboratory and field conditions, but the results were conflicting. To evaluate the effectiveness of nutrient supplementation, N sources (NO₃‐N and NH₄‐N) and P sources (PO₄‐P) were applied to the simulated diesel‐polluted seawater in the N/P ratio of 10:1 and 20:1, respectively. The results showed that the addition of nutrients increased the oil biodegradation rate and the counts of petroleum degrading bacteria (PDB) and heterotrophic bacteria (HB). A strongly positive correlation existed (the interrelated coefficient was nearly 0.9) between the percentage ratio of PDB/HB and the oil biodegradation rates, and therefore the percentage ratio of PDB/HB could be used as a good indicator to predict oil biodegradation. Among the four samples treated with nutrients, the biodegradation efficiency of the group amended with NO₃‐N and PO₄‐P in the ratio of 10:1 (10NO₃‐P group) was as much as 75.8 %, while in the 10NH₄‐P, 20NO₃‐P and 20NH₄‐P groups this value was 61.3 %, 52.4 % and 40.5, respectively. It would take natural degradation without nutrient supplementation about 78 days to achieve the result obtained within 14 days with 10NO₃‐P amendment . Chemical and microbiological analyses confirmed that the addition of nutrients in the same N/P ratio remarkably enhanced the biodegradation rate and the counts of microorganisms in the NO₃‐N treated groups, indicating that the microorganisms tend to utilize NO₃‐N rather than NH₄‐N as their growth N source. When the same kind of N source was added to the system, the promoted efficiency in the 10:1 (N/P ratio) groups was notable compared to the 20:1 groups, i.e., adding nutrients in the ratio of 10:1 is superior in the stimulation of oil biodegradation to the ratio of 20:1.     

Morphological,physiological, and biochemical characteristics of a benzoate-degrading strain <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> 1CP under stress conditions

Ability of actinobacteria Rhodococcus opacus 1CP to survive under unfavorable conditions and retain its biodegradation activity was assessed. The morphological and ultrastructural features of R. opacus 1CP cells degrading benzoate in the presence of oxidants and stress-protecting agents were investigated. The cells of R. opacus 1CP were resistant to oxidative stress caused by up to 100 mM H₂O₂ or up to 25 μM juglone (5-oxy-1,4-naphthoquinone). After 2 h of stress impact, changes in the fatty acid composition, increased activity of antioxidant enzymes, and changes in cell morphology and ultrastructure were observed. The strain retained its ability to degrade benzoate. Quercetin had a protective effect on benzoate-degrading cells of R. opacus 1CP. The strategy for cells survival under unfavorable conditions was formulated, which included decreased cell size/volume and formation of densely-packed cell conglomerates, in which the cells are embedded into a common matrix. Formation of conglomerates may probably be considered as a means for protecting the cells against aggressive environmental factors. The multicellular conglomerate structure and the matrix material impede the penetration of toxic substances into the conglomerates, promoting survival of the cells located inside.     

Studies on Non-Covalent Interaction of Coumarin Attached Pyrimidine and 1-Methyl Indole 1,2,3 Triazole Analogues with Intermolecular Telomeric G-Quadruplex DNA Using ESI-MS and Spectroscopy

In the present study, electrospray ionization mass spectrometry (ESI-MS) and spectroscopy have been used to evaluate the non-covalent interaction, stoichiometry, and selectivity of two synthetic coumarin-attached nucleoside and non-nucleoside 1,2,3-triazoles, namely, (1-(5-(hydroxymethyl)-4-(4-((2-oxo-2H-chromen-4-yloxy)methyl)-1H-1,2,3-triazol-1-yl)tetrahydro-furan-2-yl)5-methyl pyrimidine-2,4(1H,3H)-dione (Tr1) and 4-((1-((-1-methyl-1H-indol-2-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)-2H-chromen-2-one (Tr2) with two different human telomeric intermolecular G-quadruplex DNA structures formed by d(T₂AG₃) and d(T₂AG₃)₂ sequences. ESI-MS studies indicate that Tr1 specifically interacts with four-stranded intermolecular parallel quadruplex complex, whereas Tr2 interacts with two hairpin as well as four-stranded intermolecular parallel quadruplex complexes. UV–Visible spectroscopic studies suggest that Tr1 and Tr2 interact with G-quadruplex structure and unwind them. Job plots show that stoichiometry of ligand:quadruplex DNA is 1:1. Circular dichroism (CD) studies of G-quadruplex DNA and Tr1/Tr2 ligands manifest that they unfold DNA on interaction. Fluorescence studies demonstrate that ligand molecules intercalate between the two stacks of quadruplex DNA and non-radiative energy transfer occurs between the excited ligand molecules (donor) and quadruplex DNA (acceptor), resulting in enhancement of fluorescence emission intensity. Thus, these studies suggest that nucleoside and non-nucleoside ligands efficiently interact with d(T₂AG₃) and d(T₂AG₃)₂ G-quadruplex DNA but the interaction is not alike with all kinds of quadruplex DNA, this is probably due to the variation in the pharmacophores and structure of the ligand molecules.     

三七皂苷类自毒物质降解细菌分离及其降解特性

三七是我国的名贵药材,但由于连作障碍发生严重,因此土壤中自毒物质的积累成为导致三七连作障碍发生的主要原因之一。生物降解土壤中的自毒物质是缓解连作障碍的有效措施,为筛选并利用降解菌使土壤中皂苷类自毒物质快速消减,该研究以皂苷类自毒物质为筛选靶标,采用富集和驯化策略,从连作三七根际土壤中分离、筛选三七皂苷类自毒物质降解细菌,结合16S rRNA基因测序对高活性菌株进行分类鉴定,并对筛选得到的高活性菌株SC3的降解特性进行了研究。结果表明:(1)从三七根际土壤中成功分离出8株潜在自毒物质降解细菌,初筛评价结果显示SC3菌株对三七总皂苷的降解率最高,达87.42%。(2)通过16S rRNA基因序列分析,编号SC3的高活性菌株被鉴定为寡养单胞菌属(Stenotrophomonas)细菌。(3)在相同培养条件下,菌株SC3对单体皂苷Rb₁的降解效果强于对Rg₁的降解。(4)在液体培养条件下,底物浓度、接种量和培养温度均会显著影响SC3菌株对单体皂苷Rb₁的降解效果。综上表明,采用富集和驯化策略可以有效筛选自毒物质降解细菌,SC3菌株具有消除连作土壤中皂苷类自毒物质的潜力。该研究结果为连作土壤修复提供了生物资源,并为今后深入研究皂苷降解机制提供了理论依据。     

陕北地区石油污染土壤中不动杆菌属的筛选、鉴定及降解性能

从陕北地区石油污染土壤中分离鉴定得到两株不动杆菌属(Acinetobacter sp.)的高效石油降解菌A.sp 1和A.sp 2,分别从盐浓度、pH值、氮源、磷源和接种量等因素进行研究以确定其最佳石油降解条件,并进一步通过GC-MS(Gas ChromatographyMass Spectrometer)方法分析其在最佳条件下对原油组分的不同降解性能。结果显示:A.sp 1在盐浓度为1%、pH值为6—7、磷源为KH2PO4和K2HPO4、氮源为尿素和接种量为4%的条件下,最高降解率可达到60%。A.sp 2在盐浓度为1%、pH值为7—9、磷源为KH2PO4和K2HPO4、氮源为硝酸铵和接种量为8%的条件下,最高降解率可达到67%。GC-MS分析结果表明,菌株A.sp 1对石油烃类C21—C25有明显的降解效果,菌株A.sp 2对石油烃类C20—C30的降解效果较好。     

The role of sphingosine kinase 1/sphingosine-1-phosphate pathway in the myogenic tone of posterior cerebral arteries

Aims

The goal of the current study was to determine whether the sphingosine kinase 1 (SK1)/sphingosine-1-phosphate (S1P) pathway is involved in myogenic vasoconstriction under normal physiological conditions. In the present study, we assessed whether endogenous S1P generated by pressure participates in myogenic vasoconstriction and which signaling pathways are involved in SK1/S1P-induced myogenic response under normal physiological conditions.

Methods and Results

We measured pressure-induced myogenic response, Ca²⁺ concentration, and 20 kDa myosin light chain phosphorylation (MLC₂₀) in rabbit posterior cerebral arteries (PCAs). SK1 was expressed and activated by elevated transmural pressure in rabbit PCAs. Translocation of SK1 by pressure elevation was blocked in the absence of external Ca²⁺ and in the presence of mechanosensitive ion channel and voltage-sensitive Ca²⁺ channel blockers. Pressure-induced myogenic tone was inhibited in rabbit PCAs treated with sphingosine kinase inhibitor (SKI), but was augmented by treatment with NaF, which is an inhibitor of sphingosine-1-phosphate phosphohydrolase. Exogenous S1P further augmented pressure-induced myogenic responses. Pressure induced an increase in Ca²⁺ concentration leading to the development of myogenic tone, which was inhibited by SKI. Exogenous S1P further increased the pressure-induced increased Ca²⁺ concentration and myogenic tone, but SKI had no effect. Pressure- and exogenous S1P-induced myogenic tone was inhibited by pre-treatment with the Rho kinase inhibitor and NADPH oxidase inhibitors. Pressure- and exogenous S1P-induced myogenic tone were inhibited by pre-treatment with S1P receptor blockers, W146 (S1P1), JTE013 (S1P2), and CAY10444 (S1P3). MLC₂₀ phosphorylation was increased when the transmural pressure was raised from 40 to 80 mmHg and exogenous S1P further increased MLC₂₀ phosphorylation. The pressure-induced increase of MLC₂₀ phosphorylation was inhibited by pre-treatment of arteries with SKI.

Conclusions

Our results suggest that the SK1/S1P pathway may play an important role in pressure-induced myogenic responses in rabbit PCAs under normal physiological conditions.     

Natural isoflavones regulate the quadruplex–duplex competition in human telomeric DNA

Effects of natural isoflavones on the structural competition of human telomeric G-quadruplex d[AG₃(T₂AG₃)₃] and its related Watson–Crick duplex d[AG₃(T₂AG₃)₃-(C₃TA₂)₃C₃T] are investigated by using circular dichroism (CD), ESI-MS, fluorescence quenching measurement, CD stopped-flow kinetic experiment, UV spectroscopy and molecular modeling methods. It is intriguing to find out that isoflavones can stabilize the G-quadruplex structure but destabilize its corresponding Watson–Crick duplex and this discriminated interaction is intensified by molecular crowding environments. Kinetic experiments indicate that the dissociation rate of quadruplex (k_{obs290 nm}) is decreased by 40.3% at the daidzin/DNA molar ratio of 1.0 in K⁺, whereas in Na⁺ the observed rate constant is reduced by about 12.0%. Furthermore, glycosidic daidzin significantly induces a structural transition of the polymorphic G-quadruplex into the antiparallel conformation in K⁺. This is the first report on the recognition of isoflavones with conformational polymorphism of G-quadruplex, which suggests that natural isoflavone constituents potentially exhibit distinct regulation on the structural competition of quadruplex versus duplex in human telomeric DNA.     

Emulsified BMVC derivative induced filtration for G-quadruplex DNA structural separation

A novel method based on emulsion/filtration is introduced for G-quadruplex DNA structural separation. We first synthesized a lipophilic analogue of BMVC, 3,6-Bis(1-methyl-4-vinylpyridinium)-9-(12′-bromododecyl) carbazole diiodide (BMVC-12C-Br), which can form an oil-in-water (o/w) phase emulsion. Due to the binding preferences of BMVC-12C-Br emulsion to some specific DNA structures, the large emulsion (∼2 µm) bound DNA was separated from the small free DNA in the filtrate by a 0.22 µm pore size MCE membrane. This method is able to isolate the non-parallel G-quadruplexes from the parallel G-quadruplexes and the linear duplexes from both G-quadruplexes. In addition, this method allows us not only to determine the absence of the parallel G-quadruplexes of d(T₂AG₃)₄ and the presence of the parallel G-quadruplexes of d(T₂AG₃)₂ in K⁺ solution, but also to verify structural conversion from antiparallel to parallel G-quadruplexes of d[AG₃(T₂AG₃)₃] in K⁺ solution under molecular PEG condition. Moreover, this emulsion can separate the non-parallel G-quadruplexes of d(G₃CGCG₃AGGAAG₅CG₃) monomer from the parallel G-quadruplexes of its dimer in K⁺ solution. Together with NMR spectra, one can simplify the spectra for both the free DNA and the bound DNA to establish a spectrum-structure correlation for further structural analysis.     

Bacterial community profiles on feathers during composting as determined by terminal restriction fragment length polymorphism analysis of 16S rDNA genes

Composting is one of the more economical and environmentally safe methods of recycling feather waste generated by the poultry industry, since 90% of the feather weight consists of crude keratin protein, and feathers contain 15% N. However, the keratin in waste feathers is resistant to biodegradation and may require the addition of bacterial inocula to enhance the degradation process during composting. Two keratin-degrading bacteria isolated from plumage of wild songbirds and identified as Bacillus licheneformis (OWU 1411T) and Streptomyces sp. (OWU 1441) were inoculated into poultry feather composts (1.13×10⁸ cfu g^–1 feathers) and co-composted with poultry litter and straw in 200-l compost vessels. Composting temperatures, as well as CO₂ and NH₃ evolution, were measured in these vessels to determine the effects of inoculation on the rate and extent of poultry feather decomposition during composting. Terminal restriction fragment length polymorphisms of 16S rRNA genes were used to follow changes in microbial community structure during composting. The results indicated that extensive carbon conversion occurred in both treatments (55.5 and 56.1%). The addition of the bacterial inocula did not enhance the rate of waste feather composting. The microbial community structure over time was very similar in inoculated and uninoculated waste feather composts.     

Identification and mapping of a novel blackleg resistance locus LepR4 in the progenies from Brassica napus × B. rapa subsp. sylvestris

Blackleg, caused by Leptosphaeria maculans, is one of the most economically important diseases of Brassica napus worldwide. Two blackleg-resistant lines, 16S and 61446, were developed through interspecific hybridization between B. napus and B. rapa subsp. sylvestris and backcrossing to B. napus. Classical genetic analysis demonstrated that a single recessive gene in both lines conferred resistance to L. maculans and that the resistance alleles were allelic. Using BC₁ progeny derived from each resistant plant, this locus was mapped to B. napus linkage group N6 and was flanked by microsatellite markers sN2189b and sORH72a in an interval of about 10 cM, in a region equivalent to about 6 Mb of B. rapa DNA sequence. This new resistance gene locus was designated as LepR4. The two lines were evaluated for resistance to a wide range of L. maculans isolates using cotyledon inoculation tests under controlled environment conditions, and for stem canker resistance in blackleg field nurseries. Results indicated that line 16S, carrying LepR4a, was highly resistant to all isolates tested on cotyledons and had a high level of stem canker resistance under field conditions. Line 61446, carrying LepR4b, was only resistant to some of the isolates tested on cotyledons and was weakly resistant to stem canker under field conditions.     

A mammalian cellular system for the concomitant study of neoplastic transformation and somatic mutation.

A mammalian cellular system, utilizing Syrian hamster embryo cells, was developed for the concomitant study of neoplastic transformation and somatic mutation. Chemically induced somatic mutation of the cells was assayed at two genetic loci. Mutants deficient in hypoxanthine phosphoribosyl transferase (HPRT) were detected by the production of colonies resistant to 8-azaguanine (AG^r) or 6-thioguanine (TG^r) and mutants with an altered Na⁺/K⁺ ATPase were detected by the production of colonies resistant to ouabain (Oua^r). Colonies resistant to each of the three selective agents were isolated and characterized. AG^r and TG^r resistant cells maintained their resistance to the selective agent after isolation and growth in the absence of the drug, displayed a low reversion frequency, and possessed less than 1% of the HPRT activity of the wild-type cells. AG^r cells were also resistant to the cytotoxicity effects of 6TG. Oua^r cells also maintained their resistance to ouabain and were less sensitive to the inhibition of ⁸⁶Rb uptake by ouabain than the wild-type cells. The spontaneous frequency of all three types of resistant cells was <10^?6, but the mutation frequency was significantly increased following exposure of the cells to known mutagens in a dosage-dependent manner. These properties indicate that AG^r and TG^r cells posess a mutation in the structural or regulatory gene for HPRT, and that Oua^r cells have an altered Na⁺/K⁺ ATPase.The factors involved in the quantification of the mutation frequencies of hamster embryo cells following exposure to carcinogens were determined. Cytotoxicity was assayed by a reduction in the cloning efficiency of the treated cells. The recovery efficiencies of the resistant cells were measured by reconstitution experiments and the degree of cross feeding effects of HPRT^? cells was determined. The expression time of the mutations following exposure of the cells to carcinogens was also examined, and the mutation frequencie at the two loci of hamster embryo cells following exposure to MNNG or benzo(a)-pyrene (B(a)P) were determined. Employing this system, a quantitative comparison can be made between the frequencies of somatic mutation and morphological transformation.     

Association of two SNPs in the coding region of the insulin-like growth factor 1 receptor (IGF1R) gene with growth-related traits in Angus cattle

The insulin-like growth factor 1 (IGF-1) is considered to be a factor that mainly regulates growth, differentiation, and the maintenance of various function in numerous tissues through binding to a family of transmembrane tyrosine kinase receptors, signaling primarily through the insulin-like growth factor 1 receptor (IGF-1R) encoded by the IGF1R gene. The objectives of the present study were to estimate the allele and genotype frequencies of the IGF1R/MspI (silent mutation within exon 12) and the IGF1R/TaqI (within the 3′ untranslated region, 3′UTR) gene polymorphisms in beef cattle and to determine associations between these polymorphisms and growth traits. In a preliminary study on 310 Angus calves, association analyses with three production traits (birth weight, BWT; weaning weight adjusted to 210 days, WWT₂₁₀; and average daily gain, ADG) were conducted. The GG genotype of the IGF1R/e12/MspI polymorphism was significantly associated (P?≤?0.05) with a higher WWT₂₁₀ (+5.06 kg) compared to the AG genotype. Polymorphism within the 3′UTR had no significant effect on growth traits. The effect of combined genotypes was also examined. At WWT₂₁₀, calves with the GG/AA and GG/AG combinations were heavier than calves with the AG/AA and AG/AG combined genotypes (P?≤?0.05). To our knowledge, this is the first report of a polymorphism within the coding region of the Bos taurus IGF1R gene.