| Pinus tabulaeformis f. shekannesis, a Pinus taxon, is morphologically similar with P. tabulaeformis. It only distributes in the upper part of Damaijiegou, Heshangyuan forest farm, Fuxian county, Shaanxi province with pure forest area of 337.3 ha. Compared with P. tabulaeformis, P. tabulaeformis f. shekannesis has many advantages, such as tall and straight trunk, well natural pruning, well natural reproduction ability, barren resistance, strong adaptability, large timber production per unit area, etc. P. tabulaeformis f. shekannesis is known as an excellent tree to build ecological forest and fast-grown forest in the loess plateau region. Therefore, there should be a positive protection and reasonable development and utilization for Pinus tabulaeformis f. shekannesis. However, there still is a controversy about the taxonomic status of P. tabulaeformis f. sekannesis in Pinus genus, and is also lack of understanding about the present situation of its genetic diversity. All of these, thus, restrict the systemic research and the effective protection of this excellent genetic resource. This study selected P. tabulaeformis f. sekannesis and its 5 nearest wild relatives(P. tabulaeformis, P. tabulaeformis var. mukdensis, P. tabulaeformis var. mukdensis, P. massoniana, and P. henryi) to assess the taxonomic status of P. tabulaeformis f. shekanensis. We studied the taxonomic status of P. tabulaeformis f. shekannesis on the basis of molecular taxonomy, morphological taxonomy, anatomical taxonomy and chemotaxonomy. Moreover, genetic diversity of two only extant populations of P. tabulaeformis f. shekannesis was investigated by using morphology characteristics and molecular markers. The main results are included as follows:1. Thirteen SSR primer pairs were screened from 82 primers pairs which were developed from other Pinus genus. These 13 primers had high polymorphism, good specificity and stability. A total of 153 individuals come from six Pinus taxa were screened by these 13 primer pairs with results of 84 allelic variation, each locus of allelic variation of range from 3 to 15, with an average of 6.50, polymorphism information content(PIC) from 0.293 to 0.875, with a mean of 0.600. Population genetic analysis(neighbor-joining cluster analysis, principal coordinate analysis, and structure clustering) revealed that P. tabulaeformis f. shekanensis and P. tabulaeformis were conspecific(P. tabulaeformis).2. Cluster analysis and principal component analysis were produced according to the morphological characteristics, wood anatomical characteristics, oleoresin characteristics and essential oil characteristics data in six Pinus taxa. Clustering analysis results, based on the mean of each index in the six taxa, revealed that P. tabulaeformis f. shekannesis and P. tabulaeformis will always be clustered together, the remaining 4 taxa then clustered with former two in different order. According to the raw data of each index to each individual, results of cluster analysis and PCA mapping showed that 153 individual desultorily mixed with each other and the relationship was very complicated. These results revealed there was no specificity of morphological characteristics, wood anatomical characteristics, oleoresin characteristics and essential oil characteristics among the six taxa.3. Single factor analysis of variance was performed between P. tabulaeformis f. shekannesis and P. tabulaeformis in morphological characteristics, wood anatomical characteristics, oleoresin characteristics and essential oil characteristics. It showed that there were several indexes showed the extremely significant or significant differences between the two taxa, which were needle size and shape, size and shape of male cone, size and shape of the cone, weight of the cones, seed scale length, seed wing size, seed size weight, the angle between branch and trunk, the size of artificial form-quotient, the size of resin canals in the stem, the size of the tracheid lumen diameter, the size of the tracheid wall cavity ratio, the width of the earlywood and latewood, 12 chemical composition content of the oleoresin(3-Carene, No. 35 unknown diterpene, Palustric acid, Abietic acid, 7,13,15-Abietatrienoic acid, No. 56 unknown diterpene, No. 58 unknown diterpene, ?-Pinene, Isopimaric acid, Isopalustrate acid, Neoabietic acid, 45 unknown diterpene), 9 chemical composition content of the essential oil(?-Caryophellene, ?-Muurolene, ?-Pinene, Spathulenol, Eudesmol, Ledol, Globulol, ?-Cadinol, Cembrene). Meanwhile, according to a report in the previous studies, we found a number of significant or extremely significant differences between the two taxa, which were the shape and crack of bark, the colour of needle, the angle between lateral branch and trunk(the same as the this study), cone shape and size(the same as the this study), stem straightness, wood stiffness, natural pruning condition, resin canals number and size in the stem, pith ray number in the stem, the size of vascular bundles in the needles, the size and proportion of endothelial layer in the needles, resin canals number and size in the needles, fat cells number of resin canals in the needles, the size of pollen, the size of corpus of pollen, the size of airsacs of pollen and the width of aperture of pollen. Therefore, according to the results of SSR cluster analysis, all kinds of stable differences characteristics between P. tabulaeformis f. shekannesis and P. tabulaeformis, and combined with the concept of form, variety and subspecies, it can be concluded that P. tabulaeformis f. shekanensis should be classified as a variety of P. tabuliformis. In addition, in this paper, according to the taxonomic status of P. tabulaeformis f. shekannesis(P. tabulaeformis variety), the characteristics of the P. tabulaeformis f. shekannesis forestry, and combined with relevant historical events, the origin of P. tabulaeformis f. shekannesis was discussed.4. In this research, 17 phenotypic traits in 100 trees from 2 existing populations in P. tabulaeformis f. shekannesis were selected and investigated. The seed wings only had significant differences between the populations, the rest of the 16 phenotypic traits between/within populations were highly significant differences. Variation coefficients(CV) of phenotypic traits in P. tabulaeformis f. shekannesis were 11.34 %( needles), 12.80 %( cones), 14.06 %( seed scale), 19.47%(seed wing) and 30.16 %( seed), respectively, which also indicated that P. tabulaeformis f. shekannesis has a higher levels of a variety of phenotypic traits. The average of phenotypic differentiation coefficient(Vst) of the 17 phenotypic traits was 45.03%, revealing 45.03% phenotypic variation was exist to between populations and 54.97% within populations. Most of 17 phenotypic traits in P. tabulaeformis f. shekannesis presented significant or highly significant positive correlation, of which needle length, needle width, cone length, cone diameter, cone dry weight, seed scale length and seed wing length could be regarded of important phenotypic traits when we do some related research. In total, 74 allelic variations were detected by 13 SSR primer pairs, each locus of allelic variation of range from 3 to 10, with an average of 5.69, polymorphism information content from 0.415 to 0.865, with a mean of 0.629. At the species level, the Nei's genetic diversity index was 0.3956 and the Shannon information index was 1.0021. At the population level, both Nei's genetic diversity index and Shannon information index indicated that the genetic diversity of population 1(sunny-slope) is slightly larger than population 2(shady-slope). Coefficient of gene differentiation(Gst) between 2 populations of P. tabulaeformis f. shekannesis was 0.0232 and Shannon's population differentiation coefficient((Isp- pop)/Isp) between 2 populations was 0.0270, which revealed that there was less than 3% of the genetic variation existed between populations and more than 97% of genetic variation within the population. Results of molecular variance(AMOVA) were consistent with the coefficient of gene differentiation(Gst) and the group of differentiation coefficient((Isp- pop)/Isp), which also pointed out that 2.67% of genetic variation existed between populations and 97.33% of genetic variation within population. Finally, based on the current situation of genetic diversity in P. tabulaeformis f. shekannesis and combined with the characteristics of P. tabulaeformis f. shekannesis forestry, some protection strategies were proposed. |
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