| The tea[Camellia sinensis (L.) O. Kuntze] breeding and resources preservation depend on the fully understanding of the genetic variation amount, distribution and evolution of Gene Tank. It is difficult to study the tea's inherited disorders and evolution by traditional measures such as form and agronomic characters analysis, while molecular marker is an effective method in studying the genetic diversity and genetic differentiation. In this study, genetic diversity in twenty-five local cultivars of the tea plants in Guangdong province and five contrastive cultivars from different regions were assessed using morphological, SRAP, AFLP and ISSR makers for the first time, and the genetic relationship among the accessions was revealed by cluster analysis at various levels. The main results are as follows:1. Analysis on the 10 morphological characters of the 30 tea cultivars showed that the average coefficient variation was 32.15%, the biggest was the fine-hair (42.41%) , and the smallest was growth period of tea leaves (18.52%) . The cluster analysis of morphological characters showed that the 30 tea cultivars could be divided into 4 groups. There were 17 in the first group, 10 in the second group, and the third group contained two contrastive cultivars: Yunnan-dayezhong, and Lingyun-baimaocha, there was only one group-Hainan-dayezhong in the forth group.2. In this study, genetic diversity in tea plant was assessed using SRAP makers for the first time at home and abroad, which showed that SRAP was an effective, cheap and dependable marker for polymorphism analysis in tea plant. A total of 127 amplified fragments and 114 polymorphic fragments were detected by applying 21 SRAP primer combinations. The polymorphic fragment percentage was 88.67%. The number of amplified fragments and polymorphic fragments per primer combination were 6.05 and 5.43 respectively. 30 tea accessions were divided into four groups of A, B and C at genetic distance of 0.388, 83.33% of which were classified into group A. Group A could also be divided into I, II and IIIsub-groups at genetic distance of 0.314. There were 13 in the I subgroup, 2 cultivars in the II subgroup and 10 in the III subgroup.3. The AFLP silver staining protocol was used for amplification reaction in the experiment, twenty-five local cultivars of tea plant in Guangdong province and five contrastive cultivars were tested and analyzed using 5 primer combinations, 401 bands were obtained, 338 bands (84.3%) of them were found to be polymorphic,80.2 bands were obtained by every one primer pair on the average. The amplification fragment ranged from 50bp to 580bp. Among the 30 cultivars, the greatest genetic distances was between Guibei-daye and Fenghuang-shuixian (0.6075) , the lowest genetic distance was between Qingliangshan-cha and Qingyuan-bijiacha (0.1275) . It was reported here that the local tea plant of Guangdong province was characterized by a high genetic diversity. A dendrogram showing genetic relationships between them was constructed by an unweighted pair-group method with arithmetical averages (UPGMA) of SPSS 12.0. 30 cultivars could be divided into three groups according to the combine line 0.40 of the genetic distance. This result shows that local Guangdong tea plants' consanguineous relationship is diversified and concentrative.4. The 15 ISSR primers generated 104 polymorphic bands in the 30 cultivars, 101 bands (96.89%) of which were found to be polymorphic,6.93 bands were obtained by every oneprimer pair on the average. Among the 30 cultivars the average genetic distances were 0.5018 ,the greatest genetic distances was between Pingyuan-guoducha and others (0.7600) . We find 30 cultivars can be divided into three groups according to the combine line 0.6500 of the genetic distance which containe 18,10 and 2 cultivars respectively. The 18 cultivars can be grouped to A type(60.00%), according to the combine line 0.5500 of the genetic distance. A type can be divided into I, II subgroups, the I subgroup containes 14 cultivars and the II subgroup containes 4.5. There was remarkable correlation between the genetic distance of AFLP and ISSR (r =0.3089, P<0.01) molecular markers, significant correlation between SRAP and ISSR (r =0.1577, P< 0.05) , while the result of SRAP and AFLP had no significant correlation (r =-0.0975, P > 0.05). Based on the gray relational and analytical method, a relational level among SRAP, AFLP and ISSR makers and morphological cluster dendrogram was studied. The following conclusions were drawn. The relational level of SRAP marker lied in the fist place, the relational level of ISSR marker was in the second, and the relational level of AFLP marker was in the third.6. By analyzing the integrated data from SRAP,AFLP and ISSR data, they were classified into 6 groups, according to the combine line 0.40 of the genetic distance. The first group included 3 cultivars, those were Guangzhou-xiaoyebaixin, Guangzhou-xiaoyeqingxin and Pingyuan-guoducha, the second group contained 9 cultivars, those were Luoding-xiaoye, Xingning-guantiancha, Guangning-dayeqingxin, Puning-xiaoye, Guangning-qingguidaye, Gaozhou-baixin, Lechang-kucha, Guibei-dayezhong and Jiangxi-ningzhouzhong, the third group included 4 cultivars, those were Qingliangshan-cha, Fengshun-matucha, Liannan-daye, Dongyuan-shangguancha, Lechang-baimaocha, Renhua-baimaocha, Ruyuan-baimaocha, Taishan-baiyuncha, Qingyuan-bijiacha, Wuhuatianzhushan-cha and Nankunshan-baimaocha, the forth group included 4 cultivars, those were Yangchun-baimaocha, Huiyang-xiaoye, Yunnan-dayezhong and Hainan-dayezhong, the fifth group was Lingyun-baimaocha, the sixth group included Fenghuang-shuixian and Shiguping-wulong. The results reflected the genetic relationship among tea accessions more rationally than using different markers separately. This told us that different methods could not be replaced simply and it was feasible and proper to integrate various data.In summary, the quantitative criteria for evaluating genetic diversity and sampling strategy for genetic resources conservation and utilization of local cultivars of Guangdong province tea plant had been provided, and the results could have been used to guide genetic resources collection, conservation and elite breed selection of tea plant. This study was significant for genetic diversity research and conservation of local cultivars of tea plant in Guangdong province. |
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