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中国科学院昆明植物研究所知识管理系统
Knowledge Management System of Kunming Institute of Botany,CAS
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GST,p < 0.001) and low levels of seed-based gene flow. C. debaoensis (Cycadaceae) is an endangered species restricted to the border of Guangxi and Yunnan province in southwest China. This species has been classified into two types: sand and karst, according to the soil matrix they grow on. We examined chloroplast sequence variation of the cpDNA sequences from 11 populations of this species. Significant population genetic differentiation was detected (GST= 0.684 and FST = 0.74160). There was marked genetic differentiation between populations in the sand and karst regions and no expansion was detected. Climate changes during glacial periods have had significant effects on the current distribution of cycads. The molecular phylogenetic data, together with the geographic distribution of the haplotypes, suggest that C. debaoensis experienced range contraction during glacial periods, and that the current populations are still confined to the original refugia in southwest China which have favorable habitats in glacial period. These results imply that small refugia were maintained in both sand and karst regions during the LGM (last glacial maximum). This species had no postglacial recolonization and only stayed in these refugia up to now. The low within-population diversity of C. debaoensis suggests that there were strong bottleneck events or founder effects within each separate region during the Quaternary climatic oscillations. Relatively high genetic and haplotype diversities were detected in the newly discovered populations, which located at intermediate locality of sand regions and had morphological variation; this is probably the consequence of the admixture of different haplotypes colonizing the area from separate sources. C. micholitzii occurs in the Annan Highlands in central Vietnam near the Laos border. C. bifida occurs in North Vietnam; its distribution extends across the border into adjacent localities in Guangxi and Yunnan in China. For the comparability between them,theywere considered as the same species C. micholitzii by many academicians. The cpDNA sequences from 11 populations showed that these very controversial species, C. micholitzii and C. bifida, is paraphyletic and should belong to the same species C. micholitzii. AMOVA analysis showed that the component of among-population within region/species (76.46%) was unexpectedly larger than the among-species/region component (14.97%), which also indicates that there is no justification for recognizing two species as C. micholitzii and C. bifida. This hypothesis was also supported by the geological data, especially the neotectonic history of the indo-china block, which started to move south since Oligocene and cause the geographic isolation of these two groups. Therefore, the most likely explanation to the phenotypic similarities between these two groups may be the retention of ancestral polymorphisms in the paraphyletic group due to incomplete lineage sorting. Furthermore, the similarities may also be ascribed to pollen-mediated gene flow among geographically proximate populations and/or phenotypic convergence under similar selection schemes in the same region. C.micholitzi had the higest genetic diversity (HT = 0.980,) and genetic differentiation (GST = 0.830, NST = 0.915) among the C. micholitzii complex. The high genetic diversity might be attributed to its long evolutionary history, highly diverse habitats. The ineffective mode of seed dispersal and dramatic neotectonic movement in the distribution range of this species could result in the high genetic differentiation. 2. Phylogeographic analysis based on nuclear ribosomal sequences, We sequenced the nrDNA ITS in all 27 populations sampled, 7 haplotypes were identified, among which C. micholitzii had 6, while C. multipinnata, C. longipetiolula and C. debaoensis shared the remaining one. Compared to chloroplast genes, nuclear genes had higher correlation between genetic and geographical distance, but lower interspecies differentiation (54.42% vs 25.24%). Phylogeographical structure of C. micholitzii and C.bifida based on ITS Variation was consistent with the morphology differentiation. This similar in nuclear gene should be ascribed to pollen-mediated gene flow among geographically proximate populations.Long-distance gene flow over the two groups was clearly interrupted, which brought on the nrDNA genetic differenciation between the geographically isolated groups, to a certain extent affected the morphological variation. 3. Interspecies relationships among Cycas micholitzii complex, We analysed chloroplast sequence variation of the atpB-rbcL and psbA-trnH intergenic spacers in 27 populations sampled of C. micholitzii complex, AMOVA analysis showed that the component of among-species/region component (59.21%). However, phylogenic analysis showed that the haplotypes of C. micholitzii complex couldn`t grouped into four clusters closely corresponding to the narrowly defined C. micholitzi, C. multipinnata, C. debaoensis and C. longipetiolula. We concluded that the conflict may result from several factors: firstly incomplete lineage sorting of C. micholitzii; secondly hybridization/introgression of sympatrically cycads, which would be supported by evidence base on nrDNA ITS sequences; thirdly intramolecular recombination in cpDNA of cycads; eventually the neotectonic movement in the distribution range of this 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study can reveal important biological features of plants and answers to a certain degree in phylogeny and distribution of genetic materials and so forth. By hard working of cytologists, chromosome data of plants have been increased to a great abundance, but yet disorderly distributed in different magazines, which made researches based on the whole chromosome data of one taxon rarely launched. Scientific databases have become increasingly indispensable as researching data growing daily. As Cytological studies are booming in China, in order to fill the absence of digital and statistical data of plant chromosome researches and chromosome atlas, we started to develop a Chinese Seed Plants Chromosome Database, aiming to construct a database and start to record published chromosome data of Chinese seed plants. Based on this database, we chose the part of gymnosperms and gave a discussion to the features of its chromosomes’ evolution and variation. Cytological experiments have been applied to some important phyto-groups for phylogeny research and germplasm identification.Part I: The Chinese Seed Plants Chromosome Database and Discussion on the features of Gymnosperms chromosomes，1 The Chinese Seed Plants Chromosome Database，The frame of database was constructed by Microsoft Access 2003. 19 items of data were included in, they are: Chinese and Latin names of family, genus and species; plant pictures, mitosis metaphase and karyotype figures; morphological characteristics and distributions of the plant; chromosome numbers and basic numbers; karyotype formula; karyotype description; origin of the plant material; literature and the source of photos. In this database, data can be checked and shared easily by extracted out in species sorted interface or family sorted interface. 120 species in 29 genera and 10 families of Gymnospers have been collected and input to the database. In Angiosperms, 61 species in 10 genera of family Magnoliaceae and 80 species in 3 genera of family Theaceae have been collected and input to the database.2 Discussion on the features of evolution and variation of Gymnosperms chromosomes，By data collection of the database, we analyzed chromosome features of the group Gymnosperm. Plants of Gymnosperm had been through a long historical evolution on earth, fossil records of which originated from the late Devonian period. Once an authoritative and major classification level in the plant kingdom, most Gymnosperms have been extinct unless conifers, cycads, Ginkgo and Getales. Three main features of Gymnosperm chromosomes are: relatively large chromosome, which can be recognized from figures in the database; constant chromosome numbers, in most families of Gymnosperm the basic chromosome number keeps a certain value; comparatively low variation, karyotype under family level differs a little. The variation of chromosomes in Gymnosperm is dominated by Robertsonian changes. Contrary to common variation type in Angiosperms, the variation from high unsymmetric karyotype to low unsymmetric karyotype was found in existence in Gymnosperm.Part II: cytology research on some important phyto-groups，3 Karyomorphology of three species in the order Huerteales and their phylogenetic implications，The karyomorphology of three species in Dipentodon (Dipentodontaceae), Perrottetia (Celastraceae), and Tapiscia (Tapisciaceae), namely Dipentodon sinicus, Perrottetia racemosa, and Tapiscia sinensis, was investigated in the study. Recent molecular research has discovered close relationships among these three genera, which has led to the establishment of the order Huerteales with Perrottetia being placed in Dipentodontaceae. Herein we report the chromosome numbers of D. sinicus and P. racemosa for the first time, and present their karyotype formulas as 2n = 34 = 22sm + 12st (D. sinicus), 2n = 20 = 11m + 9sm (P. racemosa), and 2n = 30 = 22m(2SAT) + 8sm (T. sinensis). Asymmetry of their karyotypes is categorized to be Type 3B in D. sinicus, Type 2A in P. racemosa, and Type 2A in T. sinensis. Each of the species shows special cytological features. Compared with Perrottetia, Dipentodon has a different basic chromosome number, a higher karyotype asymmetry, and different karyomorphology of its interphase nuclei, mitotic prophase, and metaphase. Thus, on the basis of these results, we have reservations regarding the suggestion of placing Dipentodon and Perrottetia together in the family Dipentodontaceae.4 Genomic analyses of intergeneric hybrids between Michelia crassipes and M. calcicola by GISH，Genomic in situ hybridization (GISH) is becoming the method of choice for identifying parental chromosomes in interspecific hybrids. Interspecific F1 hybrid between Michelia crassipes and M. calcicola, tow highly ornamental species in Michelia of Magnolicaceae, has been analized by double-colored GISH with its parents’ genome as the probe. Research gave the results that the chromosome number of the F1 hybrid is 2n=38 as the same of species in Michelia and other genera in Magnoliaceae, the basic chromosome is x=19, the karyotype formula is 2n=38=32m+6sm, and the asymmetry of karyotype is 1B type. Based on chromosome data of Michelia in our database, the karyotype of this genus is featured mostly by metacentric chromosomes and submetacentric chromosomes. In Mechelia, the variation range of submetacentric chromosomes is 4 to 18 and of the karyotype asymmetry is 1A to 2B type. Both the karyotype and karyotype asymmetry type of F1 hybrid is among the variation range of Michelia. The figure of GISH showed that all the 38 chromosomes of F1 hybrid have crossing parental signals, and signal on the no.1 and no.7 chromosome showed differences, which proved that both the parental genome have been transmitted to and recombinated in F1 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scent is a very important character in rose breeding. However, many of 25,000 rose cultivars have no scent or weak scent. The tea scent of modern roses mainly originated from Rosa odorata (Andrews) Sweet, which is one of the most important ancestors of modern cultivated roses and the very important rose breeding resource. Due to the land expanding, habitat fragmentation and so on, R. odorata has been listed as an endangered species in ‘Chinese Plant Red Data Book—Rare and Endangered Plants’ and as the third-category endangered species in ‘Chinese Rare and Endangered Protective Plants List’. Therefore, it is urgent to protect this species and studying the conservation genetics of R. odorata is essentially important to work out a strategy of conservation.R. odorata comprises three double-petaled varieties (R. odorata var. odorata, R. odorata var. erubescens, and R. odorata var. pseudindica) and one single-petaled variety (R. odorata var. gigantea). The taxonomy of the three double-petaled varieties of R. odorata has been disputed for a long time. They have been treated as intraspecific taxa of R. odorata var. gigantea or R. chinensis by different botanist. According to the morphological analyses, Hurst (1941) inferred that R. odorata var. odorata was the hybrid between R. odorata var. gigantea and R. chinensis. Therefore, in order to clarify the right protective units, two single-copy nuclear genes (GAPDH and ncpGS), together with two plastid loci (trnL-F and psbA-trnH) were applied to study the hybrid origin of the three double-petaled varieties and to identify their possible parents. Our data suggested the hybrid origin of the three double-petaled varieties. We inferred that R. odorata var. gigantea could be the maternal parent and R. chinensis cultivars be the paternal parent. It is strongly suggested that the conservation of R. odorata is the conservation of its wild type, R. odorata var. gigantea. We first applied seven microsatellite loci (SSR) coupled with a single-copy nuclear gene GAPDH to study the genetic diversity and genetic structure of R. odorata var. gigantea. The main results are shown as follows:1. Genetic diversity：R. odorata var. gigantea maintains high degree of genetic diversity within and among populations (SSR: HT = 0.738, HS = 0.569, AR = 5.583, PPB = 97.35%, I = 1.703; GAPDH: HT = 0.739, HS = 0.540). We inferred that, outcrossing, long-lived tree species, clonal reproduction and general intraspecies hybridization between individuals, have contributed to the high degree of genetic diversity in R. odorata var. gigantea.2. Genetic differentiation and genetic structure：There was some degree of genetic differentiation among populations (SSR: GST = 0.229, FST = 0.240; GAPDH: GST = 0.269). The geographic isolation limited the dispersal of pollen or seeds, which resulted in the limitation of gene flow (Nm = 0.792). Then, the limited gene flow should be accounted for the genetic differentiation. Both the results of SSR data and haplotype analysis of GAPDH indicated that, the studied populations were divided into two distinct groups by Honghe River. These two groups showed significant genetic differentiation and represented two separate evolutionary lineages, which should be recognized as two evolutionary significant units (ESUs) for conservation concerns.3. Conservation of R. odorata：R. odorata var. gigantea has been listed in the ‘National Key Protective Wild Species List (II)’. Therefore, the conservation of this species is urgent. We inferred that, the main endangered reasons should be the habitat fragmentation and the reduction of populations and individuals per population resulted from environmental damage and human activities. We proposed that the strategy of in-situ conservation combining with ex-situ conservation should be carried 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