| Pinus yunnanensis Franch. is a subtropical pine endemic to Southwest China with characteristics of light-loving, drought-resistant, poor-site tolerant, great adaptability and extensive values of timber, etc. It plays a crucial role in regional economic development and ecological restoration. At present the low, crooked and twisted individuals were appeared in P. yunnanensis forests, and the excellent resources are reducing. Even there appeared varieties (P. yunnanensis var. pygmaea) and forma (crooked P. yunnanensis). The different opinions were proposed about the phenotypic degradation of P. yunnanensis. The knowledge of the extent and pattern of intra-specific variation, formation of the genetic variation of P. yunnanensis, and the genetic relationships between P. yunnanensis and varieties or forma will be of great significance both for the collection and conversation of germplasm resources, population reconstruction and for the decision of genetic improvement strategies. In present paper 20 populations of P. yunnanensis from main distribution regions, nine populations from different altitudes were sampled in Yunnan Province of China, and varieties, forma as well as three sympatric pine species were also collected. The level and distribution of intra-specific variation, the population differentiation based on the phenotypic characters such as needle and cone, and SSR loci were evaluated to determine the proportion of variation which existed among and within populations, and to explore correlations between genetic variation and geographic or climatic parameters. The main results were as follows.(1) There were significant differences (p< 0.01) both among populations and within populations in each needle and cone trait. There existed wide and abundant genetic variation in P. yunnanensis populations. The morphological variation was more abundant in low-altitude population than middle-or high-altitude populations. But there was no difference among populations from different altitudes. The coefficients of variation (CV) was highest in the ratio of needle length to fascicle sheath length (NL/FSL), and lowest in fascicle width (FW) and needle length (NL), which indicated that FW and NL were not easily influenced by environment. There was high CV in the southern populations when compared on others. Mean number of observed alleles, effective alleles, observed heterozygosity, expected heterozygosity and Shannon’s information index were 3.7,2.0,0.470,0.429 and 0.787 per population, respectively. P. yunnanensis exhibited a medium level of genetic diversity among Pinus species. The southern populations also showed the higher genetic diversity than others. Although there were significantly morphological differences among populations, no evident differences were detected based on SSR data.(2) Most genetic diversity was found within populations, and overall genetic differentiation was low. The phenotypic differentiation coefficients among populations varied from 31.07% to 81.81% with a mean of 49.42% for needle traits, and from 5.85% to 33.51% with an average of 17.41% for cone traits. Analysis of molecular variance (AMOVA) revealed higher genetic variation within populations than among populations based on SSR data, which were also confirmed by the weak genetic differentiation (Fst=0.052). The low differentiation could further explain that the genetic variation was mainly within populations, which was correlated with phenotypic differentiation coefficients. The results were reinforced using the Bayesian software STRUCTURE, which identified 20 natural populations may be sourced from three original genetic populations, and two southern populations (YJ and SJ) were clearly separated from the others. But no clear genetic structure was found among populations. The inbreeding coefficient of each population was not significantly different from zero. P. yunnanensis was a highly outcrossing species. A low level of inter-population genetic differentiation and a high estimate of gene flow (7.525) were appeared. The strong gene flow may be attributed to the biological characteristics and the continuous distribution in a large area of this species, which weaken the genetic differentiation among populations.(3) The needle traits and genetic diversity parameters based on SSR loci of the regional populations were correlated with geographical, climatic and soil parameters. Those populations with rich genetic diversity existed in low latitude, low altitude, warm and moist environment. There were weak correlations between cone traits and environmental parameters, and no definite geographic pattern was found. The needle traits and genetic diversity parameters based on SSR loci of the altitudinal populations were not correlated with soil parameters and altitude difference. Euclidean distance estimated for phenotypic data and Nei’s (1983) genetic distance estimated for SSR data were positively correlated in P. yunnanensis populations, which suggested the two approaches were generating the same pattern of genetic diversity. There was non-significant Mantel test of the relationship between matrices of genetic distance and matrices of geographical distance, which indicated that there was no evident sign of isolation by distance (IBD). But the isolation of ecology (IBE) was more obvious than IBD. The genetic differentiation among populations may be influenced by geographical, climatic or soil factors.(4) There were no significant genetic differentiations between P. yunnanensis and P. yunnanensis var. pygmaea, crooked P. yunnanensis or P. yunnanensis var. tenuifolia based on SSR data. The genetic variation analysis of the varieties (P. yunnanensis var. pygmaea and P. yunnanensis var. tenuifolia), forma (crooked P. yunnanensis) and other sympatric pine species (P. densata, P. kesiya var. langbianensis and P. massoniana) of P. yunnanensis showed that high genetic diversity in P. kesiya var. langbianensis population and low genetic diversity in P. yunnanensis var. tenuifolia. Those varieties, forma and species were classified into three main groups by UPGMA cluster analysis based on Nei’s (1983) genetic distance. Among them five out of seven species or varieties were clustered into one group. P. kesiya var. langbianensis was near to this group. P. massoniana was the further to this group. The most similar ones were found between P. yunnanensis and crooked P. yunnanensis, then grouped with P. yunnanensis var. pygmaea. Also P. yunnanensis var. tenuifolia and P. densata showed a high similarity coefficient. The UPGMA analysis based on SSR data revealed a really close genetic distance between P. yunnanensis, crooked P. yunnanensis, P. yunnanensis var. pygmaea, P. yunnanensis var. tenuifolia, which suggested those varieties or forma belonged to a same species and the phenotypic degeneration may be caused by the negative selection or environmental degradation.(5) The populations that should be given priority for conservation were proposed by focusing on the maintenance of genetic diversity. Generally because of environmental factors on the genetic differentiation future germplasm should be considered to conserve in various regions with geographical and climactic differences. The southern populations should be a focus of in situ conservation efforts due to their higher level of genetic diversity. For this study, majority of the total genetic variation was within populations and minority among populations, which suggested that large numbers of samples from few populations would be expected to contain most of the neutral genetic diversity. The superior populations should have the priority of in situ conservation as a whole. It also would be available ex situ conservation for excellent individuals selected from the superior populations. In order to extend the reforestation range or afford material for adaptability studies, it should be considered to conserve the populations in extreme conditions such as high latitude or altitude, etc. Generally because of the knowledge of low genetic differentiation, high phenotypic difference and the negative selection or environmental degradation, the genetic improvement of forest stand with good site conditions depending on natural regeneration could be realized by keeping excellent plants and logging low, crooked as well as twisted plants, and the genetic improvement of forest stand with poor site conditions could be gradually renewed and regenerated by using excellent varieties of P. yunnanensis combined with cultivation measures such as soil improvement and water collection, etc, which could restore the dominant status of main plant population, optimize the ecological function and provide efficient use of resources. |
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