We investigated the hierarchical genetic structure of SSR (simple sequence repeats) and cpDNA (chloroplast DNA) polymorphisms among and within populations of Primula sieboldii, a heterostylous clonal herb. Seven out of eight populations at the study site, located in a mountainous region of Nagano Prefecture, had each developed alongside a different stream, and the other occurred on a flat area 70 m from the nearest stream. The magnitude of genetic differentiation among streamside populations in maternally inherited cpDNA (Phi = 0.341) was much higher than that in biparentally inherited SSRs (Phi = 0.011). This result suggests that seed dispersal among streams was restricted, and pollen was the primary agent of gene flow among streamside populations. In contrast, genetic differentiation among subpopulations within streams were low at both markers (Phi = 0.053 for cpDNA, Phi = 0.025 for SSR). This low differentiation among subpopulations in cpDNA compared with that among streamside populations suggest that seed dispersal occur along the stream probably during flooding. This hypothesis was supported by the fact that in cpDNA haplotypes, no clear genetic structure was detected within the streamside population, while a significant genetic structure was found within 20 m in the nonstreamside population. Furthermore, within the streamside populations, two pairs of ramets with identical multilocus genotypes for eight SSR loci were distantly (> 50 m) distributed along the same streamside, suggesting dispersal of clonal propagule. Our study showed that the heterogeneity of the landscape can influence gene flow and hence spatial genetic structure.

To evaluate the effects of flowering phenology on pollen flow distance and spatial genetic structure in a population of a bumblebeepollinated herb, Primula sieboldii, we investigated the flowering phenology of 1712 flowers of 97 genets in a population in Nagano Prefecture, Japan, and constructed a mating model based on the observed mating pattern, which was revealed by paternity analysis using 11 microsatellite markers. The effects of flowering phenology were inferred by comparing estimated pollen flow distance and the level of heterozygosity in the next generation between two scenarios. In the first scenario, both the intergenet distance and flowering phenology influenced mating opportunity, while in the second scenario only intergenet distance influenced mating opportunity. Although the frequency distribution of pollen flow distance at the population level did not differ significantly between the two scenarios, the mean pollen flow distance of several flowers increased by more than 10 m as a result of variation in flowering phenology. Furthermore, accounting for flowering phenology predicted change in heterozygosity in the next generation from Ϫ0.04 to 0.07. The results showed that flowering phenology can affect pollen flow distance and spatial genetic structure.Key words: bumblebee; flowering time; heterostyly; mating model; microsatellite; Nagano Prefecture; paternity analysis; Primulaceae.Knowledge of effective pollen flow distance has great importance in evolutionary and conservation genetics, because pollen flow distance can strongly affect spatial genetic structure and hence the spatial scale of local adaptation and the total level of genetic diversity in a population (Loveless and Hamrick, 1984). Many theoretical and simulation studies have shown that homozygous patches develop rapidly due to inbreeding and genetic drift when both seed and pollen flow are spatially restricted within a population (Wright, 1943;Turner et al., 1982;Sokal and Wartenberg, 1983;Ohsawa et al., 1993). However, most of these studies assumed that there is no variation in flowering phenology (i.e., flowering time and the number of flowers) among plants within a population. In reality, large variations in flowering phenology are often observed among plants within a population (Pors and Werner, 1989;Dieringer, 1991;Washitani et al., 1991;Okayama et al., 2003). These variations in flowering phenology may induce spatial and temporal variation in flower density and affect the degree of spatial restriction of pollen flow; researchers have shown that flower density can strongly affect pollen flow distance (Levin and Kerster, 1969 population of insect-pollinated American mahogany (Swietenia humilis Zucc.). Likewise, mass (synchronous) flowering of neighboring plants may increase the proportion of pollen flow between neighbors and hence may shorten the mean pollen flow distance. The effect of flowering phenology on pollen flow distance may have significant consequences for spatial genetic structure (Schmitt, 1983;Loveless and Hamrick, 1984...

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To promote programs for the conservation and restoration of the endangered species Primula sieboldii, we examined genetic variation at eight microsatellite loci among and within 32 remnant wild populations throughout Japan. Total allelic diversity within a population was higher in larger populations, but not so after rarefaction adjustment. The positive relationship between population size and the inbreeding coefficient may suggest that more heterozygous genets tend to survive the habitat contraction possibly because of the higher fitness associated with heterozygosity. By principal coordinate analysis and Bayesian analysis, we detected four genetic groups (Hokkaido, northern Honshu, central Honshu, and western Japan), which could be recognized as management units of P. sieboldii. If supplementation with plants from other populations were planned, it should be conducted among populations which belong to the same management unit and which are likely to represent the same adaptive variation.

Primula sieboldii (E. Morren), the ancestor of the Japanese garden Primula, is in danger of extinction in the wild. Genetic diversity is a key component for conservation efforts associated with population management. Genetic diversity in visible traits and several molecular makers were evaluated, respectively. Since it is difficult to determine the degree to which genetic and environmental differences contribute to observed phenotypic variation in natural habitats. A common-garden approach is used. The contribution of genetic and environmental differences to variation in petal shape and area of Primula sieboldii were evaluated. Samples from 108 genets gathered from five natural populations in three regions of Japan were analyzed in a commongarden experiment and also analyzed using eight microsatellite markers. From the results of quantitative evaluation based on image analysis, broad genetic variation in petal traits within populations and low level of population differentiation was found. For all petal shapes, Q st was smaller than F st , suggesting that wild populations might be under moderate selective pressure for a specific phenotype. For petal area, Q st was nearly equal to F st , suggesting that population differentiation has been caused mainly by genetic drift.