Clonality is a common phenomenon in plants, allowing genets to persist asexually for much longer periods of time than ramets. The relative frequency of sexual vs. asexual reproduction determines long-term dominance and persistence of clonal plants at the landscape scale. One of the most familiar and valued clonal plants in North America is aspen (Populus tremuloides). Previous researchers have suggested that aspen in xeric landscapes of the intermountain west represent genets of great chronological age, maintained via clonal expansion in the near absence of sexual reproduction. We synthesized microsatellite data from 1371 ramets in two large sampling grids in Utah. We found a surprisingly large number of distinct genets, some covering large spatial areas, but most represented by only one to a few individual ramets at a sampling scale of 50 m. In general, multi-ramet genets were spatially cohesive, although some genets appear to be fragmented remnants of much larger clones. We conclude that recent sexual reproduction in these landscapes is a stronger contributor to standing genetic variation at the population level than the accumulation of somatic mutations, and that even some of the spatially large clones may not be as ancient as previously supposed. Further, a striking majority of the largest genets in both study areas had three alleles at one or more loci, suggesting triploidy or aneuploidy. These genets tended to be spatially clustered but not closely related. Together, these findings substantially advance our understanding of clonal dynamics in western North American aspen, and set the stage for a broad range of future studies.

Microsatellite data are widely used to test ecological and evolutionary hypotheses in wild populations. In this paper, we consider three typical sources of scoring errors capable of biasing biological conclusions: stuttering, large-allele dropout and null alleles. We describe methods to detect errors and propose conventions to mitigate scoring errors and report error rates in studies of wild populations. Finally, we discuss potential bias in ecological or evolutionary conclusions based on data sets containing these scoring errors. AbstractMicrosatellite data are widely used to test ecological and evolutionary hypotheses in wild populations. In this paper, we consider three typical sources of scoring errors capable of biasing biological conclusions: stuttering, large-allele dropout and null alleles. We describe methods to detect errors and propose conventions to mitigate scoring errors and report error rates in studies of wild populations. Finally, we discuss potential bias in ecological or evolutionary conclusions based on data sets containing these scoring errors.

We analyzed mitochondrial (mt) DNA restriction fragment length polymorphisms (RFLPs) associated with cytochrome oxidase, subunit I (coxI)-related gene sequences in 268 trees derived from 19 natural populations of three species of pines from California (USA): Monterey pine (Pinus radiata D. Don), bishop pine (P. Muricata D. Don), and knobcone pine (P. attenuata Lemm.). Total genomic DNA was digested with four restriction endonucleases and probed with a 750-bp fragment of the mitochondrialcoxI gene amplified fromP. attenuata via the polymerase chain reaction (PCR). ThecoxI gene is repeated at least 4 times in some populations, and all variants that we observed resulted from complex rearrangements rather than from point mutations. There was limited intrapopulation variation, but strong differentiation among populations. When applied to haplotype frequencies, Nei's gene diversity within populations (Hs) averaged 7% (±3), and Gst varied from 75% forP. Radiata to 96% forP. muricata. The high degree of population differentiation for mtDNA suggests that it can be a powerful marker of population differences, but its rapid rate of structural evolution appears to result from recombination among a limited number of repetitive elements-giving frequent homoplasious fragment phenotypes. The phylogenetic trees disagreed with results from chloroplast DNA, nuclear gene, and morphological studies.

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We determined the DNA sequence of a 2.7-kb cpDNA XbaI fragment from douglas-fir [Pseudotsuga menziesii (Mirb.) Franco]. RFLPs revealed by the 2.7-kb XbaI clone were observed to vary up to 1 kb among species within the genus Pseudotsuga and up to 200 bp among trees of P. menziesii. The polymerase chain reaction (PCR) allowed the locus of polymorphism to be identified, and the variable region was then sequenced in a second Douglas-fir tree, a single tree of a related species, Japanese Douglas-fir (P. japonica), and in a species lacking a mutation hotspot in the region, Pinus radiata (Monterey pine). The locus of polymorphism is characterized by hundreds of base pairs of imperfect, tandem direct repeats flanked by a partially duplicated and an intact trn Y-GUA gene. The duplication is direct in orientation and consists of 43 bp of the 3' end of trnY and 25 bp of its 3' flanking sequence. Tandem repeats show high sequence similarity to a 27-bp region of the trnY gene that overlaps one end of the duplication. The two trees of Douglas-fir sequenced differed by a single tandem repeat unit, whereas these trees differed from the Japanese Douglas-fir sequenced by approximately 34 repeat units. Repetitive DNA in the Pseudotsuga cpDNA hotspot was most likely generated at the time of the partial trnY gene duplication and these sequences expanded by slipped-strand mispairing and unequal crossing-over.

Eleven highly polymorphic microsatellite markers were used to determine the genetic structure and levels of diversity in 51 natural populations of Pinus oocarpa across its geographic range of 3000 km in Mesoamerica. The study also included 17 populations of Pinus patula and Pinus tecunumanii chosen for their resistance or susceptibility to the pitch canker fungus based on previous research. Seedlings from all 68 populations were screened for pitch canker resistance, and results were correlated to mean genetic diversity and collection site variables. Results indicate that P. oocarpa exhibits average to above-average levels of genetic diversity (A ¼ 19:82, A R ¼ 11:86, H E ¼ 0:711) relative to other conifers. Most populations were out of Hardy-Weinberg equilibrium, and a high degree of inbreeding was found in the species (F IS ¼ 0:150). Bayesian analysis grouped P. oocarpa into four genetic clusters highly correlated to geography and distinct from P. patula and P. tecunumanii. Historic gene flow across P. oocarpa clusters was observed (N m ¼ 1:1-2:7), but the most pronounced values were found between P. oocarpa and P. tecunumanii (low-altitude provenances) in Central America (N m ¼ 9:7). Pinus oocarpa appears to have two main centers of diversity, one in the Eje Transversal Volcánico in central Mexico and the other in Central America. Introgression between P. oocarpa and P. tecunumanii populations appears to be common. Pinus oocarpa populations showed high resistance to pitch canker (stemkill 3%-8%), a disease that the species has presumably coevolved with in Mesoamerica. Resistance was significantly correlated to the latitude, longitude, and altitude of the collection site but not to any genetic-diversity parameters or degree of admixture with P. tecunumanii.

Ponderosa pine (Pinus ponderosa) is among the most broadly distributed conifer species of western North America, where it possesses considerable ecological, esthetic, and commercial value. It exhibits complicated patterns of morphological and genetic variation, suggesting that it may be in the process of differentiating into distinct regional lineages. A robust analysis of genetic variation across the ponderosa pine complex is necessary to ensure the effectiveness of management and conservation efforts given the species' large distribution, the existence of many isolated disjunct populations, and the potential susceptibility of some populations to climate change and other threats. We used highly polymorphic nuclear microsatellite markers and isozyme markers from 3113 trees in 104 populations to assess genetic variation and structure across the geographic range of ponderosa pine. The results reveal pervasive inbreeding and patterns of genetic diversity consistent with the hypothesis that ponderosa existed in small, as-yet-undetected Pleistocene glacial refugia north of southern Arizona and New Mexico. The substructuring of genetic variation within the species complex was consistent with its division into two varieties, with genetic clusters within varieties generally associated with latitudinal zones. The analyses indicate widespread gene flow and/or recent common ancestry among genetic clusters within varieties, but not between varieties. Isolated disjunct populations had lower genetic variation by some measures and greater genetic differentiation than main-range populations. These results should be useful for decision-making and conservation planning related to this widespread and important species.

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