Cloning,heterologous expression,and characterization of recombinant class II cytochromes c from Rhodopseudomonas palustris

The cytochrome (cyt) c', cyt c(556), and cyt c(2) genes from Rhodopseudomonas palustris have been cloned; recombinant cyt c' and cyt c(556) have been expressed, purified, and characterized. Unlike mitochondrial cyt c, these two proteins are structurally similar to cyt b(562), in which the heme is embedded in a four-helix bundle. The hemes in both recombinant proteins form covalent thioether links to two Cys residues. UV/vis spectra of the Fe(II) and Fe(III) states of the recombinant cyts are identical with those of the corresponding native proteins. Equilibrium unfolding measurements in guanidine hydrochloride solutions confirm that native Fe(II)-cyt c(556) is more stable than the corresponding state of Fe(III)-cyt c(556) (DeltaDeltaG(f)(o) =22 kJ/mol).     

Molecular cloning and functional expression of soybean allene oxide synthases

A plant allene oxide synthase (AOS) reacting with 13S-hydroperoxy-9Z,11E,15Z-octadecatrienoic acid (13-HPOT), a lipoxygenase product of alpha-linolenic acid, provides an allene oxide which functions as an intermediate for jasmonic acid (JA) synthesis, making AOS a key enzyme regulating the JA level in plants. Although AOSs in various plants have been investigated, there is only limited information about AOSs in soybean (Glycine max). In this study, we cloned and characterized two soybean AOSs, GmAOS1 and GmAOS2, sharing 95% homology in the predicted amino acid sequences. GmAOS1 and GmAOS2 were composed of 564 and 559 amino acids respectively, with predicted N-terminal chloroplast-targeting signal peptides. Both AOSs expressed in Escherichia coli were selective for 13S-hydroperoxides of alpha-linolenic and linoleic acids, suggesting the potential of GmAOS1 and GmAOS2 to contribute to JA synthesis. GmAOS1 and GmAOS2 were expressed in leaves, stems, and roots, suggesting broad distribution in a soybean plant.     

Phylogeography and ecological niche modeling reveal evolutionary history of Leiolepis ocellata (Squamata,Leiolepidae)

Leiolepis ocellata is a lizard species distributing in topographically diverse habitats in northern Thailand. To explore its evolutionary history, 113 samples of L. ocellata were collected from 11 localities covering its distributional range in northern Thailand, and sequenced for mtDNA fragments (Cyt b and ND2). Pairwise comparisons across sampling localities yielded significant genetic differentiation (F _ST and Jost''s D) but no clear pattern of isolation by distance could be demonstrated based on the Mantel test. Phylogenetic and network analyses highlighted six haplogroups. Their divergence times were estimated to occur during the Pleistocene, much more recent than major orogenic events affecting northern Thailand. Instead, the results suggested that lineage divergences, of particularly eastern and western haplogroups of the region, coincided with the major rivers in the region (Yom river and Ping river, respectively), indicating vicariance in response to riverine barriers. Furthermore, ecological niche modeling suggested an expansion of suitable habitats of L. ocellata, when LGM‐liked conditions. This expansion potentially facilitated their dispersal among adjacent localities leading to lineage diversification and genetic admixture, after the riverine divergence.     

Highly specific and sensitive detection of Burkholderia pseudomallei genomic DNA by CRISPR-Cas12a

Detection of Burkholderia pseudomallei, a causative bacterium for melioidosis, remains a challenging undertaking due to long assay time, laboratory requirements, and the lack of specificity and sensitivity of many current assays. In this study, we are presenting a novel method that circumvents those issues by utilizing CRISPR-Cas12a coupled with isothermal amplification to identify B. pseudomallei DNA from clinical isolates. Through in silico search for conserved CRISPR-Cas12a target sites, we engineered the CRISPR-Cas12a to contain a highly specific spacer to B. pseudomallei, named crBP34. The crBP34-based detection assay can detect as few as 40 copies of B. pseudomallei genomic DNA while discriminating against other tested common pathogens. When coupled with a lateral flow dipstick, the assay readout can be simply performed without the loss of sensitivity and does not require expensive equipment. This crBP34-based detection assay provides high sensitivity, specificity and simple detection method for B. pseudomallei DNA. Direct use of this assay on clinical samples may require further optimization as these samples are complexed with high level of human DNA.     

Isolation and characterization of polymorphic microsatellite loci in the raccoon (Procyon lotor)

We report the isolation and characterization of 17 polymorphic microsatellite loci in the North American raccoon (Procyon lotor). These loci exhibit high levels of allelic diversity, with between four and 13 alleles per locus, and heterozygosity, with observed values of 0.500-1.000 in a sample of 20 individuals. All genotypes conformed to Hardy-Weinberg expectations and there were no instances of linkage disequilibrium detected.     

Tests of proportional hazards and proportional odds models for grouped survival data

In this paper, we derive score test statistics to discriminate between proportional hazards and proportional odds models for grouped survival data. These models are embedded within a power family transformation in order to obtain the score tests. In simple cases, some small-sample results are obtained for the score statistics using Monte Carlo simulations. Score statistics have distributions well approximated by the chi-squared distribution. Real examples illustrate the proposed tests.     

Environmental assessment of energy production from municipal solid waste incineration

Background, Aims and Scope During the combustion of municipal solid waste (MSW), energy is produced which can be utilized to generate electricity. However, electricity production from incineration has to be evaluated from the point view of the environmental performance. In this study, environmental impacts of electricity production from waste incineration plant in Thailand are compared with those from Thai conventional power plants. Methods The evaluation is based on a life cycle perspective using life cycle assessment (LCA) as the evaluation tool. Since MSW incineration provides two services, viz., waste management and electricity production, the conventional power production system is expanded to include landfilling without energy recovery, which is the most commonly used waste management system in Thailand, to provide the equivalent function of waste management. Results The study shows that the incineration performs better than conventional power plants vis-à-vis global warming and photochemical ozone formation, but not for acidification and nutrient enrichment. Discussion There are some aspects which may influence this result. If landfilling with gas collection and flaring systems is included in the analysis along with conventional power production instead of landfilling without energy recovery, the expanded system could become more favorable than the incineration in the global warming point of view. In addition, if the installation of deNO_x process is employed in the MSW incineration process, nitrogen dioxide can be reduced with a consequent reduction of acidification and nutrient enrichment potentials. However, the conventional power plants still have lower acidification and nutrient enrichment potentials. Conclusions The study shows that incineration could not play the major role for electricity production, but in addition to being a waste management option, could be considered as a complement to conventional power production. To promote incineration as a benign waste management option, appropriate deNO_x and dioxin removal processes should be provided. Separation of high moisture content waste fractions from the waste to be incinerated and improvement of the operation efficiency of the incineration plant must be considered to improve the environmental performance of MSW incineration. Recommendations This study provides an overall picture and impacts, and hence, can support a decision-making process for implementation of MSW incineration. The results obtained in this study could provide valuable information to implement incineration. But it should be noted that the results show the characteristics only from some viewpoints. Outlook Further analysis is required to evaluate the electricity production of the incineration plant from other environmental aspects such as toxicity and land-use.     

Genetic analysis of hybridization between white-handed (Hylobates lar) and pileated (Hylobates pileatus) gibbons in a contact zone in Khao Yai National Park,Thailand

Primates - Natural hybridization has played various roles in the evolutionary history of primates. Its consequences range from genetic introgression between taxa, formation of hybrid zones, and...