Isolation and characterization of microsatellite markers in hoary marmots (Marmota caligata)

Microsatellite loci were developed from hoary marmots (Marmota caligata) to aid in the investigation of the social structure and mating system of this species. Seven of the microsatellite loci developed were found to be moderately polymorphic with between two and seven alleles per locus. In addition to the microsatellites developed in hoary marmots we also tested markers developed for other scuirids, namely European alpine marmots (M. marmota), Columbian ground squirrels (Spermophilus columbianus) and European ground squirrels (S. citellus). Of these markers, 13 were polymorphic when amplified in hoary marmots with between two and nine alleles per locus.     

gerud1.0: a computer program for the reconstruction of parental genotypes from progeny arrays using multilocus DNA data

We have no standard computer algorithm for the reconstruction of parental genotypes from the data generated by molecular studies of progeny arrays. Here I present a computer program that uses the multilocus genotypes of parents and offspring to reconstruct the genotypes of unknown parents contributing gametes to a progeny array for which one parent is known a priori. A second program performs simulations to assess the reliability of the algorithm under various scenarios. These programs will aid scientists engaged in parentage analyses, particularly in species with large clutches.     

Genetic variation in two populations of the rough‐skinned newt (Taricha granulosa) assessed using novel tetranucleotide microsatellite loci

Seven novel tetranucleotide microsatellite loci were identified from a partial genomic DNA library, enriched for GATA‐motif microsatellites, from the rough‐skinned newt (Taricha granulosa). All loci were polymorphic, and one displayed a high frequency null allele. A related species, T. rivularis, displays strong site fidelity and detectable population structure over small spatial scales, so we assessed genetic variation in two samples of T. granulosa separated by 16 km. Distributions of allele frequencies differ significantly between our two sites, but small values of F_ST and Rho_ST suggest that the populations are linked by a large amount of gene flow.     

Efficient resolution of parentage in dogs by amplification of microsatellites

Parentage control has been performed for 15 litters from 12 different dog breeds by amplification of microsatellites. As it was possible to include all putative parents in all cases, they were solved by exclusion. Discrimination between parents/non-parents was made after genotyping of 6–9 microsatellite loci. In 12 of the cases all but one of the alleged fathers were excluded while in three cases it was unambiguously shown that superfecundation had taken place. Furthermore, one inclusion case concerning disputed maternity has been investigated. Maternity indices were calculated for 12 loci and probability of maternity was estimated to be 99-99%. These results testify that microsatellites can be applied very efficiently for resolution of parentage in dogs.     

Development and characterization of six new microsatellite markers for the silver‐ or gold‐lipped pearl oyster,Pinctada maxima (Pteriidae)

Six di‐, tri‐ and tetranucleotide microsatellite loci were developed for the silver‐ or gold‐lipped pearl oyster Pinctada maxima using a linker‐ligated, magnetic bead enrichment protocol. Based on a minimum of 134 Indonesian pearl oyster samples, number of alleles and observed heterozygosity at each locus ranged from six to 17 alleles and from 0.172 to 0.813 (mean = 0.448), respectively. Mean polymorphic information content for the six loci was 0.562. These loci should be very useful in DNA parentage analyses and population differentiation of P. maxima in Australia and Indonesia.     

Relative accuracy of three common methods of parentage analysis in natural populations

Parentage studies and family reconstructions have become increasingly popular for investigating a range of evolutionary, ecological and behavioural processes in natural populations. However, a number of different assignment methods have emerged in common use and the accuracy of each may differ in relation to the number of loci examined, allelic diversity, incomplete sampling of all candidate parents and the presence of genotyping errors. Here, we examine how these factors affect the accuracy of three popular parentage inference methods (colony , famoz and an exclusion‐Bayes’ theorem approach by Christie (Molecular Ecology Resources, 2010a, 10, 115) to resolve true parent–offspring pairs using simulated data. Our findings demonstrate that accuracy increases with the number and diversity of loci. These were clearly the most important factors in obtaining accurate assignments explaining 75–90% of variance in overall accuracy across 60 simulated scenarios. Furthermore, the proportion of candidate parents sampled had a small but significant impact on the susceptibility of each method to either false‐positive or false‐negative assignments. Within the range of values simulated, colony outperformed FaMoz, which outperformed the exclusion‐Bayes’ theorem method. However, with 20 or more highly polymorphic loci, all methods could be applied with confidence. Our results show that for parentage inference in natural populations, careful consideration of the number and quality of markers will increase the accuracy of assignments and mitigate the effects of incomplete sampling of parental populations.     

An improvement on the maximum likelihood reconstruction of pedigrees from marker data

Many methods have been proposed to reconstruct the pedigree of a sample of individuals from their multilocus marker genotypes. These methods, like those in other fields of statistical inferences, may suffer from both type I (falsely related) and type II (falsely unrelated) errors. In sibship reconstruction, type I errors come from the spurious fusion of two or more small sibships into a single sibship, and type II errors originate from the spurious splitting of a large sibship into two or more small sibships. In this study I investigate the tendencies of both types of errors made by the likelihood methods in sibship reconstruction, using both analytical and simulation approaches. I propose an improvement on the likelihood methods to reduce sibship splitting, and thus type II errors by downscaling the number of inferred siblings sharing the same genotype at a locus. Simulations are then conducted to compare the accuracy of the original and improved likelihood methods in sibship reconstruction of a large sample of individuals in full-sib families of the same small size, the same large size and highly variable sizes, using a variable number of loci with a variable number of alleles per locus. The methods were also applied to the analysis of a salmon data set. I show that my scaling scheme prevents effectively the splitting of large sibships, and reduces type II errors greatly with little increase in type I errors. As a result, it improves the overall accuracy of sibship assignments, except when sibships are expected to be uniformly small or marker information is unrealistically scarce.     

NMπ—improved re‐implementation of NM+, a software for estimating gene dispersal and mating patterns

This study introduces the NMπ computer program designed for estimation of plant mating system and seed and pollen dispersal kernels. NMπ is a re‐implementation of the NM+ program and provides new features such as support for multicore processors, explicit treatment of dioecy, the possibility of incorporating uniparentally cytoplasmic markers, the possibility of assessing assortative mating due to phenotypic similarity and inference about offspring genealogies. The probability model of parentage (the neighbourhood model) accounts for missing data and genotyping errors, which can be estimated along with regular parameters of the mating system. The program has virtually no restrictions with respect to a number of individuals, markers or phenotypic characters. A console version of NMπ can be run under a wide variety of operating systems, including Windows, Linux or Mac OS. For Windows users, a graphical user interface is provided to facilitate operating the software. The program, user manual and example data are available on http://www.ukw.edu.pl/pracownicy/plik/igor_chybicki/3694/ .     

Change in citrus leaf lipids during freeze-thaw stress

Fatty acids in the leaves of the citrus hybrid B5-9-68, a BC₁ progeny [(Citrus paradisi x Poncirus trifoliata) x C. sinensis], were compared with those in P. trifoliata and C. sinensis. Total lipid and triacylglycerol fatty acid profiles of the cold hardened hybrid were similar to the profiles of the hardy P. trifoliata and different from the profiles of the less hardy C. sinensis. When subjected to a freeze regime, the unhardened hybrid lost 22% leaf fatty acids during the freeze stage and 13% during the thaw stage. Linolenic acid accounted for 98% of the fatty acid decrease. Degradation was greatest in phosphatidylcholine (89%), phosphatidylglycerol (83%), monogalactosyldiglyceride (79%) and digalactosyldiglyceride (50%). Phosphatidic acid increased 4-fold over the two stress stages. Total leaf fatty acids during the freeze-thaw regime increased 12% in the cold hardened hybrid. Three molecular species of triacylglycerol which were rich in linolenic acid increased in the hardened hybrid during the freeze-thaw regime. The increase in highly unsaturated triacylglycerol species under freeze-thaw stress suggests that triacylglycerol has a role in maintaining the fluidity of biomembranes during freezing conditions.     

Raman spectroscopic study of the valinomycin--KSCN complex

This paper reports the first Raman spectroscopic study of the potassium complex of the cation-specific antibiotic valinomycin. Complete Raman spectra (140 to 3600 cm^?1) of crystalline valinomycin-KSCN and its CCl₄, CHCl₃ and C₂H₅OH solutions are presented and used to probe the structure of the complex in these environments. In all cases a single, narrow peak is observed in the ester CO stretch region (1750 to 1775 cm^?1) which contrasts strongly with the broad bands observed in solutions of uncomplexed valinomycin. This is consistent with the presence of a single conformation in which all six ester CO groups co-ordinate an enclosed potassium ion. We find that although the ester CO stretch frequencies of the complex are similar in the solid state and in non-polar solution (~1770 cm^?1) they are considerably different in the presence of polar solvents (~1756 cm^?1); this may indicate that the complexed potassium ion is still free to interact with nearby solvent ions (and possibly its counterion) through gaps in the hydrophobic “shield” provided by the hydrocarbon residues of valinomycin. In contrast the amide CO frequencies of the complex (~1650 cm^?1) are solvent-independent. These groups are apparently strongly hydrogen-bonded to provide a rather rigid, compact framework for the complex conformation.