Molecular database for classifying Shorea species (Dipterocarpaceae) and techniques for checking the legitimacy of timber and wood products

The extent of tropical forest has been declining, due to over-exploitation and illegal logging activities. Large quantities of unlawfully extracted timber and other wood products have been exported, mainly to developed countries. As part of the export monitoring effort, we have developed methods for extracting and analyzing DNA from wood products, such as veneers and sawn timbers made from dipterocarps, in order to identify the species from which they originated. We have also developed a chloroplast DNA database for classifying Shorea species, which are both ecologically and commercially important canopy tree species in the forests of Southeast Asia. We are able to determine the candidate species of wood samples, based on DNA sequences and anatomical data. The methods for analyzing DNA from dipterocarp wood products may have strong deterrent effects on international trade of illegitimate dipterocarp products. However, the method for analyzing DNA from wood is not perfect for all wood products and need for more improvement, especially for plywood sample. Consequently, there may be benefits for the conservation of tropical forests in Southeast Asia. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10265-010-0348-z) contains supplementary material, which is available to authorized users

B Chromosomes Have a Functional Effect on Female Sex Determination in Lake Victoria Cichlid Fishes

The endemic cichlid fishes in Lake Victoria are a model system for speciation through adaptive radiation. Although the evolution of the sex-determination system may also play a role in speciation, little is known about the sex-determination system of Lake Victoria cichlids. To understand the evolution of the sex-determination system in these fish, we performed cytogenetic analysis in 11 cichlid species from Lake Victoria. B chromosomes, which are present in addition to standard chromosomes, were found at a high prevalence rate (85%) in these cichlids. In one species, B chromosomes were female-specific. Cross-breeding using females with and without the B chromosomes demonstrated that the presence of the B chromosomes leads to a female-biased sex ratio in this species. Although B chromosomes were believed to be selfish genetic elements with little effect on phenotype and to lack protein-coding genes, the present study provides evidence that B chromosomes have a functional effect on female sex determination. FISH analysis using a BAC clone containing B chromosome DNA suggested that the B chromosomes are derived from sex chromosomes. Determination of the nucleotide sequences of this clone (104.5 kb) revealed the presence of several protein-coding genes in the B chromosome, suggesting that B chromosomes have the potential to contain functional genes. Because some sex chromosomes in amphibians and arthropods are thought to be derived from B chromosomes, the B chromosomes in Lake Victoria cichlids may represent an evolutionary transition toward the generation of sex chromosomes

Development of EST-SSR markers from an inner bark cDNA library of Fagus crenata (Fagaceae)

Fagus crenata Blume is widely distributed throughout Japanese cool-temperate deciduous broad-leaved forests, but there are two divergent groups of populations in areas with contrasting winter climates separated by Japan’s Central Mountain Range. To facilitate investigations of adaptive genetic differentiation of the species using potentially functional genes, we have collected Expressed Sequence Tags and developed Simple Sequence Repeat markers using a cDNA library constructed from cambium and surrounding tissues. In total, 270 primer pairs were designed, and 87 of the corresponding loci showed polymorphism in 16 individuals, with 2–21 alleles per locus and expected heterozygosities ranging from 0.06 to 0.97. EST-SSR markers developed in the present study will be useful for genomic analyses of F. crenata populations

An efficient method for developing SNP markers based on EST data combined with high resolution melting (HRM) analysis

Abstract Background In order to identify single nucleotide polymorphisms (SNPs) efficiently from a species with a large genome, SNPs were identified from an expressed sequence tag (EST) database combined with High Resolution Melting (HRM) analysis. Findings A total of 574 sequence tagged sites (STSs) were generated from Cryptomeria japonica and HRM analysis was used to screen for polymorphisms in these STS markers. STSs were designed in two ways: 1) putative SNP sites were identified by comparing ESTs from specific contigs, then 226 primer pairs designed for the purpose to amplify these SNPs; 2) 348 primer pairs were randomly designed using reads from the 3' end of cDNA. HRM analysis revealed that 325 markers among eight individuals were polymorphic, and that STSs, including putative SNP sites, exhibited higher levels of polymorphism. Conclusion Our results indicate that the combination of SNP screening from an EST database combined with HRM analysis is a highly efficient way to develop SNP markers for expressed genes. This method will contribute to both genetic mapping and the identification of SNPs in non-model organisms.</p

Generation of expressed sequence tags, development of microsatellite and single nucleotide polymorphism markers in Primula sieboldii E. Morren (Primulaceae) for analysis of genetic diversity in natural and horticultural populations

Primula sieboldii is a self-incompatible ornamental plant that has been cultivated for more than 300 years in Japan. In order to increase the available genomic resources for this species, 5651 expressed sequence tags (ESTs) were generated from seedling and winter bud cDNA libraries. After clustering and assembling ESTs, 2960 unigene elements that contain an homologous sequence to an S locus-linked gene of P. vulgaris were identified. In total, 127 simple sequence repeats (SSRs) were found. The most frequent di- and tri-SSR motifs were GA (78.6%) and ATT (12.9%), respectively. Twenty SSR and 40 single nucleotide polymorphism (SNP) markers were developed. These markers were evaluated for polymorphisms using 24 individuals (Y) from a population in Yatsugatake and 14 individuals (PS) from across the distributional range of P. sieboldii in Japan. The average level of expected heterozygosity within Y and PS was 0.39 and 0.48, respectively for EST-SSR markers and 0.26 and 0.29, respectively for SNP markers. The level of polymorphism for EST-SSR markers was lower than that for genomic SSR markers developed in a previous study. We also surveyed the genetic diversity of 56 cultivars, which had lower allelic richness than the Y and PS populations