Evolutionary analysis of Pinus densata (Masters), a putative Tertiary hybrid.

Summary Restriction fragment analysis and heterologous hybridization of chloroplast (cp) DNA was used to develop species-specific markers for P. tabulaeformis, P. yunnanensis and P. massoniana. Fragment patterns created by the BclI and DraI restriction enzymes and hybridization patterns to the psbC and psbD probes were distinctive among the three species. No intraspecific variation was detected with respect to any of the cpDNA markers developed in this study. The cpDNA markers obtained were subsequently used to examine the parentage of P. densata, a putative Tertiary hybrid between P. tabulaeformis and P. yunnanensis. The analysis demonstrated for the first time that P. densata populations accommodate chloroplast genomes of P. tabulaeformis and P. yunnanensis, which strongly supports earlier suggestions of the hybrid origin of this species. It appears that P. densata represents a stabilized natural hybrid that has become adapted to high mountain environments where neither of the parental species can normally grow.     

Allozyme Diversity and Population Genetic Structure of Pinus densata Master in Northwestern Yunnan, China

We investigated the levels and patterns of genetic diversity of Pinus densata Master in Yunnan. Horizontal starch-gel electrophoresis was performed on macrogametophytes collected from nine populations in northwestern Yunnan, China. Compared with other gymnosperm species, P. densata has higher mean values for all measures of genetic diversity. Allozyme polymorphism (0.99 criterion) was 97.0% and 71.4% at the species and population levels, respectively. The average number of alleles per locus was 3.1 and 2.0 at the species and population levels. Mean expected heterozygosity was substantially higher in P. densata than average values investigated for other gymnosperms both at the population (H _ep = 0.174±0.031) and at the species (H _es = 0.190) levels. Of the total genetic variation, less than 12% was partitioned among populations (G _ST = 0.112). Our allozyme survey supports the suggestion that the observed higher diversity in P. densata may be attributed partly to its hybrid origin between two genetically distinct species, P. yunnanensis and P. tabulaeformis. In addition, we suggest that introgression would give rise to the increase in genetic diversity occurring in P. densata.     

Needle morphological evidence of the homoploid hybrid origin of Pinus densata based on analysis of artificial hybrids and the putative parents,Pinus tabuliformis and Pinus yunnanensis

Genetic analyses indicate that Pinus densata is a natural homoploid hybrid originating from Pinus tabuliformis and Pinus yunnanensis. Needle morphological and anatomical features show relative species stability and can be used to identify coniferous species. Comparative analyses of these needle characteristics and phenotypic differences between the artificial hybrids, P. densata, and parental species can be used to determine the genetic and phenotypic evolutionary consequences of natural hybridization. Twelve artificial hybrid families, the two parental species, and P. densata were seeded in a high‐altitude habitat in Linzhi, Tibet. The needles of artificial hybrids and the three pine species were collected, and 24 needle morphological and anatomical traits were analyzed. Based on these results, variations in 10 needle traits among artificial hybrid families and 22 traits among species and artificial hybrids were predicted and found to be under moderate genetic control. Nineteen needle traits in artificial hybrids were similar to those in P. densata and between the two parental species, P. tabuliformis and P. yunnanensis. The ratio of plants with three needle clusters in artificial hybrids was 22.92%, which was very similar to P. densata. The eight needle traits (needle length, the mean number of stomata in sections 2 mm in length of the convex and flat sides of the needle, mean stomatal density, mesophyll/vascular bundle area ratio, mesophyll/resin canal area ratio, mesophyll/(resin canals and vascular bundles) area ratio, vascular bundle/resin canal area ratio) relative to physiological adaptability were similar to the artificial hybrids and P. densata. The similar needle features between the artificial hybrids and P. densata could be used to verify the homoploid hybrid origin of P. densata and helps to better understand of the hybridization roles in adaptation and speciation in plants.     

Systematic position of Pinus henryi (Pinaceae) as revealed by multiple evidence

The systematic position of Henry's pine (Pinus henryi Mast.), a rare species endemic to China, is still ambiguous. In this study, the phylogenetic relationships of P. henryi and its allies (P. hwangshanensis, P. tabuliformis, P. yunnanensis and P. massoniana) were revealed by multiple markers, including chemical components, DNA sequences, chloroplast microsatellites and amplified fragment length polymorphisms. The results refuted the hypothesis that P. henryi is a subspecies of P. tabuliformis or P. massoniana and supported that it might be a distinct species close to P. hwangshanensis. Pinus tabuliformis and P. yunnanensis are sister taxa. Incomplete genomic sorting, hybridization and recent divergence may explain the controversial relationships among them. Their diversification may be ascribed to habitat adaptation followed by geographic and climate changes during the Pliocene period.     

WEAK CROSSABILITY BARRIER BUT STRONG JUVENILE SELECTION SUPPORTS ECOLOGICAL SPECIATION OF THE HYBRID PINE PINUS DENSATA ON THE TIBETAN PLATEAU

Determining how a new hybrid lineage can achieve reproductive isolation is a key to understanding the process and mechanisms of homoploid hybrid speciation. Here, we evaluated the degree and nature of reproductive isolation between the ecologically successful hybrid species Pinus densata and its parental species P. tabuliformis and P. yunnanensis. We performed interspecific crosses among the three species to assess their crossability. We then conducted reciprocal transplantation experiments to evaluate their fitness differentiation, and to examine how natural populations representing different directions of introgression differ in adaptation. The crossing experiments revealed weak genetic barriers among the species. The transplantation trials showed manifest evidence of local adaptation as the three species all performed best in their native habitats. Pinus densata populations from the western edge of its distribution have evolved a strong local adaptation to the specific habitat in that range; populations representing different directions of introgressants with the two parental species all showed fitness disadvantages in this P. densata habitat. These observations illustrate that premating isolation through selection against immigrants from other habitat types or postzygotic isolation through selection against backcrosses between the three species is strong. Thus, ecological selection in combination with endogenous components and geographic isolation has likely played a significant role in the speciation of P. densata.     

Development and characterization of chloroplast microsatellite markers for <Emphasis Type="BoldItalic">Pinus massoniana</Emphasis> and their application in <Emphasis Type="BoldItalic">Pinus</Emphasis> (Pinaceae) species

Pinus massoniana is one of the important afforestation and pioneer tree species, which is widely distribute in southern China. Chloroplast simple sequence repeat markers (cpSSRs) have been widely used in studies of tree genetics, phylogenetic and breeding. We sequenced the whole chloroplast genome sequences of P. massoniana using PCR and Sanger sequencing. A total of 71 cpSSRs were identified, among which mononucleotide repeats were predominant (70.42%). Seventeen primer pairs were developed and amplification tests were conducted with 15 P. massoniana individuals. Also, cross-species amplification tests were conducted among 15 individuals per Pinus species, including P. elliottii, P. bungeana, P. armandii, P. caribaea, P. tabulaeformis, P. taiwanensis and P. yunnanensis which revealed polymorphic information content ranging from 0.2 to 0.8 and average of haploid diversity (h) ranging from 0.29 to 0.63. In addition, the polymorphic cpSSRs were useful in distinguishing the sampled pine species, and could be powerful tool in phylogenetic studies.     

中国西南四种常见松树火适应性状比较

野火对松属植物的进化和分布有重要的影响。在与野火长期抗争中,松属植物形成了一系列性状提高在易火生境中的适合度,维持种群生存与繁衍。西南地区是中国野火高发区,作为这一地区特有松树,云南松、思茅松和高山松具有一些典型的火适应性状,如厚树皮。以这3种松树和东部常见的马尾松为研究对象,比较4种松树的火适应对策。结果表明：4种松树的火适应性状存在一定的差异。与火适应相关的13项性状主成分分析显示,4个种在空间上总体是分离的,也表现出一些重叠。其中云南松火适应生活史对策是火耐受型和火依赖型的中间类型,适应会发生林冠火的生境。高山松、思茅松和马尾松都是火耐受型,通过快速高生长、厚树皮等性状适应生境不同频度的地表火。四种松树火适应对策与分布区火险基本相符。表明野火在这一区域广泛存在,并对植物进化和森林格局有重要影响。野火在西南地区松属分布和种群维持中的生态作用应被给予足够重视。     

Empirical assessment of the reproductive fitness components of the hybrid pine Pinus densata on the Tibetan Plateau

Pinus densata is distributed on the Tibetan Plateau, where it forms extensive forests at high elevations. Genetic studies have provided evidence that P. densata originated through hybridization between P. yunnanensis and P. tabuliformis. To clarify the relationships among these pines, and assess their reproductive fitness in their respective habitats, we conducted a comparative analysis of eight cone and seed morphometric traits and six reproductive traits in them. Among the eight morphometric traits examined, six appeared to be intermediate in P. densata between those of P. yunnanensis and P. tabuliformis. There were significant differences among the three pines in all of the morphometric traits, and P. densata showed greater variability in these traits than the other two pines. In contrast to the morphometric traits, the reproductive traits (including the proportions of filled and empty seeds, ovule abortion rate, seed efficiency, meiotic abnormalities during microsporogenesis and pollen viability) differed little among the three pines, indicating that they have similar overall rates of effective pollination and fertilization in their respective natural environments. Despite their location on the high plateau, natural populations of P. densata appeared to have normal levels of reproductive success, comparable to those of the two parental species in their natural habitats. This study provides empirical data characterizing the reproductive success and adaptation of a stabilized homoploid hybrid in a novel habitat that is ecologically and spatially inaccessible to its parental species. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

A critical evaluation of reproductive barriers between closely related species using DNA markers - a case study in Pinus

Former controlled crosses between twelve Pinus montana var. rostrata (Pinus mugo complex) and eight P. sylvestris clones revealed that only two P. sylvestris had efficiently fertilised P. montana. Two species-diagnostic chloroplast DNA markers were applied to verify the species purity of the parental clones. All maternal P. montana were unambiguously confirmed to belong to the P. mugo complex at both chloroplast DNA marker loci. Six P. sylvestris clones carried the `sylvestris' haplotypes. However, the same two P. sylvestris clones that had efficiently fertilised P. montana displayed the chloroplast haplotypes diagnostic to the P. mugo complex. The patterns of highly polymorphic cpDNA microsatellite markers in parents and offspring ruled out contamination by foreign pollen. We concluded that the two clones successful in the crosses represent fertile hybrids between the two species with P. mugo as the pollen donor. Consequently, DNA markers are proposed for verifying or falsifying the success of artificial fertilisation in general. The existence of crossing barriers between the two Pinus species, meaningful to the postulated natural hybridisation and the evolution of their populations in sympatric stands, was indicated and is newly discussed.     

高山松种实性状与生殖适应性

高山松(Pinus densata)是云南松(P. yunnanensis)与油松(P. tabulaeformis)自然杂交产生的二倍体杂种,是植物同倍体杂交物种形成的典型案例。高山松分布于青藏高原东南缘,占据了两个亲本种都不能正常生长的高海拔地带。良好生殖适应性的产生是物种形成的关键,但对高山松在高海拔生境下生殖适应性的变异与杂交起源遗传背景关系的研究尚未见报道。该研究通过对来自该种分布区内6个代表居群样本、13个反映高山松生殖适应性的种实性状的分析,探讨了其生殖性状在居群水平的变异模式,揭示了具有不同遗传组成和起源历史的高山松居群的生殖特性。研究结果表明:6个研究居群在种实性状上存在显著差异;可育种鳞数、总种鳞数、总种子数、可育种鳞率和可育种鳞密度等与纬度呈显著负相关关系;球果结种率与经度和生态梯度值呈显著的正相关关系;胚珠败育率则与经度和生态梯度值呈显著的负相关关系;球果长度也呈现了与经度的显著负相关。高山松球果结种率很高,平均为74%;球果的结种数、种子长和种翅长与云南松、油松及其它松属双维管亚属多数种接近;球果长度、球果总种鳞数接近油松的报道。这些结果表明,高山松在亲本种不能正常生长、繁衍的高原环境下具备正常的生殖能力;其种实特征在居群间分化显著并呈现一定的地理梯度变化。居群间的显著差异反映了其杂种起源特性,与高山松居群的遗传背景有关,也反映了处在异质生态环境中的高山松居群具有各自的生殖生态进化趋向。     

Morphological evaluation of the Pinus kesiya complex (Pinaceae)

The main aim of this study is to compare all taxa from the Pinus kesiya complex and related P. tabuliformis using statistical, ordination and discrimination techniques focusing on ten most discriminating morphological traits. Special emphasis was placed on the recently described taxa Pinus densata subsp. tibetica and P. × naxiorum (=P. yunnanensis × P. densata). Population comparisons of the particular main geographic distribution areas of Pinus kesiya, P. densata subsp. tibetica and P. yunnanensis were also conducted separately within each mentioned taxon using the same data. Pinus densata subsp. tibetica proved to be sufficiently morphologically differentiated from subsp. densata as well as from other species of the P. kesiya complex and P. tabuliformis. The recently originating hybrid P. × naxiorum appears to be intermediate between its parents. The unique shrubby taxon P. densata var. pygmaea is clearly closer to P. densata than to P. yunnanensis, to which it has been classified in Chinese floras. Populations of four distant Pinus kesiya geographic distribution areas lack substantial differences that would support the recognition of infraspecific taxa such as subsp. insularis or var. langbianensis. Pinus yunnanensis is very similar to P. kesiya, with only one trait, leaf length, being significantly different.     

Genetic variation in populations of the morphologically and ecologically variable fern <Emphasis Type="Italic">Stegnogramma pozoi</Emphasis> subsp. <Emphasis Type="Italic">mollissima</Emphasis> (Thelypteridaceae) in Japan

In Japanese Stegnogramma pozoi subsp. mollissima (Fisher ex Kunze) K. Iwats. there is the intrasubspecific variation among rbcL sequences. Northern and southern plants are genetically differentiated for maternally inherited cpDNA. In the present study we examined allozyme polymorphisms to test the hypothesis that northern and southern plants may be separate species. Based on allozyme data, the degree of gene flow among populations was estimated to be large. The artificial crossing experiments between cpDNA haplotypes also suggested that isolation has not developed among these cpDNA haplotypes. However, interpopulation genetic differentiation in cpDNA was observed even in the small area at the foot of Mt. Hakone, and the cpDNA haplotypes appear to have different habitat preferences. Received: May 1, 2001 / Accepted: September 30, 2001     

Demography and speciation history of the homoploid hybrid pine Pinus densata on the Tibetan Plateau

Pinus densata is an ecologically successful homoploid hybrid that inhabits vast areas of heterogeneous terrain on the south‐eastern Tibetan Plateau as a result of multiple waves of colonization. Its region of origin, route of colonization onto the plateau and the directions of introgression with its parental species have previously been defined, but little is known about the isolation and divergence history of its populations. In this study, we surveyed nucleotide polymorphism over eight nuclear loci in 19 representative populations of P. densata and its parental species. Using this information and coalescence simulations, we assessed the historical changes in its population size, gene flow and divergence in time and space. The results indicate a late Miocene origin for P. densata associated with the recent uplift of south‐eastern Tibet. The subsequent differentiation between geographical regions of this species began in the late Pliocene and was induced by regional topographical changes and Pleistocene glaciations. The ancestral P. densata population had a large effective population size but the central and western populations were established by limited founders, suggesting that there were severe bottlenecks during the westward migration out of the ancestral hybrid zone. After separating from their ancestral populations, population expansion occurred in all geographical regions especially in the western range. Gene flow in P. densata was restricted to geographically neighbouring populations, resulting in significant differentiation between regional groups. The new information on the divergence and demographic history of P. densata reported herein enhances our understanding of its speciation process on the Tibetan Plateau.     

Adaptive Differentiation in Seedling Traits in a Hybrid Pine Species Complex,Pinus densata and Its Parental Species,on the Tibetan Plateau

Evidence from molecular genetics demonstrates that Pinus densata is a natural homoploid hybrid originating from the parent species Pinus tabuliformis and Pinus yunnanensis, and ecological selection may have played a role in the speciation of P. densata. However, data on differentiation in adaptive traits in the species complex are scarce. In this study, we performed a common garden test on 16 seedling traits to examine the differences between P. densata and its parental species in a high altitude environment. We found that among the 16 analyzed traits, 15 were significantly different among the species. Pinus tabuliformis had much earlier bud set and a relatively higher bud set ratio but poorer seedling growth, and P. yunnanensis had opposite responses for the same traits. P. densata had the greatest fitness with higher viability and growth rates than the parents. The relatively high genetic contribution of seedling traits among populations suggested that within each species the evolutionary background is complex. The correlations between the seedling traits of a population within a species and the environmental factors indicated different impacts of the environment on species evolution. The winter temperature is among the most important climate factors that affected the fitness of the three pine species. Our investigation provides empirical evidence on adaptive differentiation among this pine species complex at seedling stages.     

The alpine homoploid hybrid Pinus densata has greater cold photosynthesis tolerance than its progenitors

The diploid hybrid tree species Pinus densata inhabits high-altitude regions, usually at higher altitudes than its parental species, P. tabulaeformis and P. yunnanensis. Its success in alpine habitat has been hypothetically attributed to enhanced cold tolerance. We tested this hypothesis by subjecting all three species to cold stress for 33 days (12/5 °C, day/night) and measuring changes in their photosynthetic function. Significant decreases in net photosynthetic rate (P_n), stomatal conductance (g_s) and transpiration rate (E) were observed in all three species in response to cold stress. The depression of P_n was primarily attributable to stomatal closure in P. densata, but to both stomatal and non-stomatal limitations in its progenitors, as the parental species had increased intercellular CO₂ concentrations (C_i) under the stress. Significant decreases in chlorophyll fluorescence parameters were also observed in both parental species, but they varied little throughout the experiment in P. densata. Furthermore, the chlorophyll and carotenoid concentrations decreased significantly under the cold stress, while anthocyanin concentrations increased, and the anthocyanin concentration in P. densata was significantly higher than in either parental species at the end of the exposure. Finally, P. densata recovered more quickly than its parental species when the cold treatment ended. These results show that P. densata maintains a high photosynthetic rate and exhibits elevated PSII activity and anthocyanin levels under cold stress. These adaptations suggest that P. densata is more cold-tolerant than its parental species and may have been important for its colonization of high alpine habitats.     

草木灰对四种松属种子发芽和幼苗生长的影响

火干扰不仅影响森林生态系统的结构与功能,而且影响群落更新与演替。为了研究草木灰对4种松属种子萌芽和幼苗生长的影响,在湖南省株洲市黄丰桥林场采集马尾松、云南松、湿地松和火炬松种子,以马尾松次生林火烧迹地的草木灰为基质,设置对照和3个草木灰处理,进行播种试验及生长观测。结果表明:(1)草木灰处理下4个物种的种子发芽率比对照试验低,并呈现随草木灰含量的增加而降低的趋势,低草木灰含量处理马尾松种子发芽率不及对照一半,中、高草木灰含量处理发芽率更低;云南松、湿地松和火炬松种子发芽率规律类似于马尾松,而高草木灰含量处理发芽率为0。(2)培养皿播种平均发芽时间比花盆播种长,无论草木灰处理还是培养条件(培养皿或花盆),马尾松和云南松种子平均发芽时间比湿地松和火炬松短。(3)4个物种种子播种14周后,对照试验中马尾松和云南松幼苗死亡率最低,而火炬松和湿地松幼苗死亡率相对增加。(4)播种14周后不同试验处理4个物种幼苗生长差异不明显。火炬松、湿地松、云南松和马尾松幼苗的单株平均干重分别达0.038、0.031、0.027、0.024 g。各物种叶干重按降序排列依次是火炬松(0.026 g)湿地松(0.019 g)云南松(0.017 g)马尾松(0.016 g),根和茎的干重比重相当,约占对应物种幼苗叶干重的三分之一。研究表明以草木灰为培养基质,马尾松和云南松种子的发芽率较高,平均发芽时间较短,更适合作为火烧迹地植被恢复先锋树种,为火干扰后选择人工促进天然更新树种提供参考。     

玉龙雪山不同针叶树种树轮宽度年表特征及其与气候因子的关系北大核心CSCD

为揭示滇西北高原树木径向生长与气候关系的规律,该研究分别在玉龙雪山采集高山松(Pinus densata)、云南松(Pinus yunnanensis)和云南铁杉(Tsuga dumosa)年轮样本,建立3个树种树轮宽度差值年表,将年轮指数与气候因子进行响应分析和冗余分析,以明确影响玉龙雪山树木径向生长的主要气候因子,为该区域森林生态系统的管理保护提供依据,并为探讨气候变化背景下滇西北森林动态提供依据。结果表明:(1)所建立的3个树种树轮宽度差值年表具有较少的低频变化,且离散程度较低、逐年变化共性较高,可代表采样点树木的年轮特征;高山松、云南松和云南铁杉差值年表的平均敏感度依次分别为0.21、0.22和0.17,样本总体代表性分别为0.96、0.96和0.94,均高于0.85的阈值,说明年表中信息具有代表性。(2)响应分析结果显示,高山松径向生长与当年2月的降水量呈显著正相关关系;云南松径向生长与当年5月的平均气温和最高温呈显著负相关关系,与当年10月平均气温呈显著正相关关系;云南铁杉径向生长与当年1月和5月的降水、当年10月相对湿度均呈显著正相关关系,与当年5月最高气温呈显著负相关关系。(3)冗余分析结果显示,当年1月降水、2月降水、5月干旱指数对3个树种均具有促进作用,且5月干旱指数的显著相关性最强;当年10月降水的增加有利于高山松和云南铁杉的径向生长,但对云南松径向生长具有抑制作用。研究认为,当年冬季降水、5月湿润条件以及10月降水是影响玉龙雪山3个树种径向生长的重要环境因子;若干暖化趋势加剧,将不利于滇西北高原3个树种的生长,从而影响区域森林生态系统结构和功能。     

Spatial distribution of cpDNA and mtDNA haplotypes in a hybrid zone betweenPinus pumila andP. parviflora var.pentaphylla (Pinaceae)

Pinus species exhibit paternal chloroplast inheritance and maternal mitochondrial inheritance. This independent inheritance of two cytoplasmic genomes provides an exceptional environment for discriminating female (seeds) and male (pollen) components of gene flow across hybridizing species. We obtained mitochondrial genetic markers diagnostic toP. parviflora var.pentaphylla andP. pumila by PCR amplification of the intron ofnad1 on mtDNA, and examined the spatial-distribution pattern of the mtDNA haplotypes in a hybrid zone betweenP. parviflora var.pentaphylla andP. pumila in the Tanigawa Mountains of Japan. These data, in conjunction with previous information on cpDNA haplotypes and needle morphology, revealed contrastive patterns of introgression of two cytoplasmic genomes. CpDNA introgression has occurred uni-directionally fromP. parviflora var.pentaphylla toP. pumila. Conversely, mtDNA introgression has occurred in the opposite direction, fromP. pumila toP. parviflora var.pentaphylla. Levels of introgression are roughly equivalent for cpDNA and mtDNA. The contrastive spatial distribution pattern of cpDNA and mtDNA haplotypes could be caused by differential movement of seeds and pollen for interspecific genetic exchange.     

The origin of remnant forest stands of Pinus tabulaeformis in southeastern Inner Mongolia

Pinus tabulaeformis is an endemic species in northern China. The northern edge of its distribution corresponds to the northern margin of the monsoon climate. Several stands of Pinus tabulaeformis beyond its major range of distribution were found on the southeastern edge of the Inner Mongolia Plateau. Phytosociological analysis shows that the community structure, species composition and regeneration ability of Pinus trees in these stands are quite different from those within its continuous range of distribution. This paper presents palynological evidence to explain how and when these remnant communities were formed. Pinus tabulaeformis entered the study area in the mid-Holocene as the summer monsoon intensified, and declined when the summer monsoon weakened. Climatic change was the driving factor for the migration of Pinus tabulaeformis. It might be supposed that the competition between pine and oak forest during the mid-Holocene warm period also affected the immigration of pine. The remnant stands of Pinus tabulaeformis were apparently favored by the microhabitat of the sandy soils in valleys when the climate became drier. The remnant stands of Pinus tabulaeformis help stabilize the stand dunes. The protection of these stands is an important task of natural conservation in the study area.     

The phylogenetic position of the endemic flat-needle pinePinus krempfii (Pinaceae) from Vietnam, based on PCR-RFLP analysis of chloroplast DNA

Pinus krempfii is morphologically very unique as compared to otherPinus species by having flat leaf-like needles. Its taxonomic position has been problematic ever since its discovery. In this study, an attempt was made to infer the taxonomic status ofP. krempfii through restriction fragment length polymorphism analysis of 12 PCR amplified chloroplast (cp) DNA regions. Phylogenetic analysis was conducted using 10 representatives of the twoPinus subgenera:Strobus andPinus. In addition, to infer the position ofP. krempfii in Pinaceae in relation with other genera, 14 representatives of eight additional genera were included in the analysis. Our cpDNA-based results indicate that: 1)P. krempfii clearly belongs to the genusPinus. This result does not favour the creation of a new genusDucampopinus in Pinaceae for this taxon. 2) Within the genusPinus, P. krempfii is more allied with species in subgenusStrobus and differs distinctly from species in subgenusPinus. 3) Despite the similarity in certain morphological and anatomical leaf and wood characters toKeteleeria andPseudolarix, the cpDNA data do not support the hypothesis for close relationship betweenP. krempfii and these two genera.