Development and characteristics of microsatellite markers for sugi (Cryptomeria japonica D. Don) derived from microsatellite-enriched libraries

Abstract

-We have developed a series of microsatellite markers for C. japonica. First, DNA fragments including microsatellite sequences were isolated from two GA-enriched genomic libraries using magnetic beads. After eliminating redundant clones and clones in which the tandem repeats were located too close to the cloning site to allow primers to be constructed, the remaining sequences could be examined for their suitability for primer design. Primer sets were designed from each conserved sequence flanking the microsatellites. We found 1 479 unique sequences in the enriched genomic libraries, of which 962 contained a tandem repeat motif, and we have been able to design 196 primer pairs using these sequences to date. The potential of these primers to amplify single fragment, and the polymorphism of the sequences they amplify, were investigated using a panel of 28 plus trees selected from Cryptomeria plantations covering the wide distributional range of the species in Japan. Forty-two of the microsatellite markers displayed a polymorphic nature throughout this panel of 28 DNA samples. The polymorphic information coefficients (PICs) ranged from 0.156 to 0.919. There was a significant correlation, between the number of repeats and the size of the PICs, according to Kendall's τ rank correlation coefficient analyses. taxodiaceae / conifer / simple sequence repeat / enrichment / primer Résumé -Développement et caractéristiques de marqueurs microsatellites pour le sugi (Cryptomeria japonica D. Don) trouvés dans des banques microsatellites enrichies. Nous avons développé une série de marqueurs microsatellites pour Cryptomeria japonica. Dans un premier temps, des fragments d'ADN comportant des séquences microsatellites ont été isolées à partir de 2 banques de séquences génomiques enrichies en GA, grâce à l'utilisation de billes magnétiques. Puis, après avoir éliminé les clones redondants et les clones pour lesquels les séquences en tandem étaient trop proches du site de clonage pour permettre aux amorces d'être construites, les séquences restantes ont été examinées afin de determiner si elles convenaient pour la construction d'amorces. Des jeux d'amorces ont été conçus à partir de chaque séquence conservée flanquant les microsattelites. Nous avons trouvé 1479 séquences uniques dans les banques génomiques enrichies, parmi lesquelles 962 contenaient 1 motif répété en tandem, et à ce jour, nous avons pu concevoir 196 paires d'amorces en utilisant ces séquences. Les capacités de ces amorces à amplifier un fragment unique, ainsi que le polymorphisme des séquences que nous avons amplifiées, ont été étudiées à partir d'un échantillon de 28 arbres « plus » sélectionnés à partir de plantations de Cryptomeria couvrant la totalité de l'aire de répartition au Japon. Quarante-deux de ces marqueurs microsatellites se sont révélés polymorphes au sein de cet échantillon. Les coefficients d'information sur le polymorphisme (PIC : Polymorphism Information Coefficient) varient de 0.156 à 0.919. Les analyses de coefficients de corrélation de ...

Gene flow and mating system in five Cryptomeria japonica D. Don seed orchards as revealed by analysis of microsatellite markers

“…We selected microsatellite markers that have shown high stability, high degrees of polymorphism, and codominant segregation in a three-generation pedigree of C. japonica (Table 2) [18,30]. The multipaternity exclusion probability [33] for the selected markers was high (>0.999) in all seed orchards.…”

Microsatellite markers reveal high allelic variation in natural populations of Cryptomeria japonica near refugial areas of the last glacial period

“…The genotypes of 11 microsatellite loci (Tani et al 2003(Tani et al , 2004Moriguchi et al 2003) in each individual were determined. One of these 11 microsatellite loci (Cjs0201) was newly developed as a marker from a microsatellite-enriched library (Tani et al 2004), and its sequence has been submitted to the DNA Data Bank of Japan (DDBJ). Of the 11 markers, 3 (Cjs0333, Cjs0520, and CS2169) were mapped to different linkage groups (Tani et al 2003), but the locations of the other markers have not yet been mapped.…”

Factors Influencing Male Reproductive Success in a Cryptomeria japonica Seed Orchard Revealed by Microsatellite Marker Analysis

“…We used six microsatellite markers that showed high stability and polymorphism ( Table 1; MORIGUCHI et al, 2003 andTANI et al, 2004) to determine the pollen donors and pollen contamination rates of seeds from nine mother trees. These markers have shown co-dominant segregation in a three-generation pedigree of C. japonica.…”