【作者】 张敏；

【导师】 陈贵林；

【作者基本信息】 内蒙古大学 ， 生物学， 2020， 硕士

【摘要】 赤芍作为传统的中药材,需求量大。近年来由于过度采挖导致野生资源匮乏,栽培赤芍成为解决市场供需的重要举措。随着赤芍栽培面积的扩大,在栽培区域出现种质混乱和相似品种混淆的现象。此外,栽培地区的环境因素对赤芍品质的影响也不可忽视。因此,开展赤芍种群遗传多样性及赤芍有效成分与气候因子的相关性研究,对建立赤芍野生资源保护区和加强赤芍品种选育以及赤芍生产区划的制定具有重要意义。本研究利用ISSR和RAPD分子标记对来自内蒙古自治区、黑龙江省和吉林省等地的28个赤芍种群以及安徽省的1个白芍种群进行遗传变异和亲缘关系的研究。同时测定了收集于内蒙古自治区、黑龙江省、河北省等地区共计22份赤芍样品的芍药苷含量,并结合30年的年均降水和年日照时数等气候因素,初步探究赤芍有效成分与气候因子的相关性。研究结果如下:1、采用14个ISSR和14个RAPD引物对赤芍基因组DNA扩增,扩增出的总条带数分别为257,215;多态性条带分别为251,209;多态性比率分别为97.8%,97.2%。2、两种分子标记共同显示野生赤芍种群的遗传多样性比栽培种群高,其中DL种群(内蒙古自治区锡林郭勒盟多伦县)的遗传多样性最高,可以作为野生赤芍保护区的候选点;此外,建议将呼伦贝尔市也作为野生资源保护区的备选区域。遗传分化参数均表明,野生赤芍种群的遗传分化主要在种群内,栽培赤芍种群的遗传分化主要在种群间。3、ISSR和RAPD的聚类分析结果均显示将29个种群分为两类,一类是赤芍种群,另一类是白芍种群,与主坐标分析结果一致,该结果可以作为鉴定赤芍和白芍的分子依据。两种分子标记共同表明,赤芍种群聚为5个亚类,其中ELC3(内蒙古自治区呼伦贝尔市鄂伦春自治旗大杨树镇)种群是独立聚类的,该种群复杂的遗传背景,需要进一步研究。4、通过UPLC测定22份赤芍样品的芍药苷含量,野生赤芍的芍药苷含量为4.9%-7.2%;而栽培赤芍的芍药苷含量为2.8%-6.8%;其中来自内蒙古自治区赤峰市翁牛特旗和黑龙江省牡丹江市林口县的样品芍药苷含量较高分别为5.4%和6.8%。相对于本研究中其他地区的赤芍样品,内蒙古自治区赤峰市翁牛特旗和黑龙江省牡丹江市林口县的赤芍质量较好。赤芍饮片的芍药苷含量为3.4%-4.9%。其本品和饮片的芍药苷含量均到达到《中华人民共和国药典》(2015版)规定值,分别为1.8%和1.5%。5、通过对赤芍有效成分与气候因子相关性分析可知:芍药苷含量与经纬度和日照时数呈正相关,与年均降水和海拔呈负相关。内蒙古自治区锡林郭勒盟南部、赤峰市中南部以及呼伦贝尔市东南部等地区日照时数长,昼夜温差大,降雨量偏少是适合赤芍生长的地区,因此可以作为内蒙古地区赤芍适宜种植基地的备选点。更多还原

【Abstract】 Paeoniae Radix Rubra is a traditional Chinese medicinal material with large demand.In recent years,due to over-excavation leading to lack of wild resources,cultivation of Paeoniae Radix Rubra has become an important measure to solve market supply and demand.With the expansion of the cultivation area of Paeoniae Radix Rubra,there is a phenomenon of confusion of germplasm and confusion of similar varieties in the cultivation area.Therefore,it is of great significance to carry out research on the genetic diversity of the Paeoniae Radix Rubra population,the correlation between the effective components of the Paeoniae Radix Rubra and the climatic factors,to establish the wild Paeoniae Radix Rubra resource protection area,strengthen the breeding of the Paeoniae Radix Rubra and formulate the Paeoniae Radix Rubra production division.In this study,ISSR and RAPD molecular markers were used to study the genetic variation and relationship of 28 Paeoniae Radix Rubra populations from Inner Mongolia Autonomous Region,Heilongjiang Province and Jilin Province and one Paeoniae Radix Alba population in Anhui Province.At the same time,the content of paeoniflorin in a total of 22 Paeoniae Radix Rubra samples collected in Inner Mongolia Autonomous Region,Heilongjiang Province,Hebei Province and other regions was measured.Combining with 30 years of annual precipitation and annual sunshine hours and other climatic factors,preliminary study on the correlation between the effective components of Radix Paeoniae Rubra and climate factors.The research results are as follows:1.Using 14 ISSR and 14 RAPD primers to amplify the Paeoniae Radix Rubra genomic DNA,the total number of amplified bands were 257,215;polymorphic bands were 251,209;polymorphism ratio were 97.8%,97.2%,respectively.2.The two molecular markers show that the genetic diversity of wild Paeoniae Radix Rubra population is higher than that of cultivated population.Among them,the genetic diversity of DL population(Duolun County,Xilinguole League,Inner Mongolia Autonomous Region)is the highest,which can be used as a candidate for wild Paeoniae Radix Rubra protection area;It is suggested that the city of Hulunbeier also be an alternative area for the protection of wild resources.The genetic differentiation parameters all indicate that the genetic differentiation of wild Paeoniae Radix Rubra population is mainly within the population,and the genetic differentiation of cultivated Paeoniae Radix Rubra population is mainly among the population.3.The cluster analysis results of ISSR and RAPD show that 29 populations are divided into two categories,one is the Paeoniae Radix Rubra population,and the other is the Paeoniae Radix Alba population,which is consistent with the principal coordination analysis results.This result can be used as the molecular basis for identifying Paeoniae Radix Rubra and Paeoniae Radix Alba.The two molecular markers together indicate that the Paeoniae Radix Rubra population is clust.ered into 5 subcategories,and the ELC3(Dayangshu Town,Elunchun Autonomous Banner,Hulunbeier City,Inner Mongolia Autonomous Region)population is independently clustered.The complex genetic background of this population requires further study.4.The content of paeoniflorin in 22 Paeoniae Radix Rubra samples was determined by UPLC.The content of paeoniflorin in wild Paeoniae Radix Rubra is 4.9%-7.2%;while the content of paeoniflorin in cultivated Paeoniae Radix Rubra is 2.8%-6.8%;The content of paeoniflorin in samples from Wengniute Banner,Chifeng City,Inner Mongolia Autonomous Region and Linkou County,Mudanjiang City,Heilongjiang Province was 5.4% and 6.8%,respectively.Compared with the Paeoniae Radix Rubra samples in other areas in this study,the quality of Paeoniae Radix Rubra in Wengniute Banner,Chifeng City,Inner Mongolia Autonomous Region and Linkou County,Mudanjiang City,Heilongjiang Province are better.The content of paeoniflorin in Paeoniae Radix Rubra cuts is 3.4%-4.9%.The content of paeoniflorin in this product and cuts reached the prescribed value of Pharmacopoeia of the People’s Republic of China(2015 edition),which were 1.8% and 1.5%,respectively.5.According to the correlation analysis of the effective components of Paeoniae Radix Rubra and climatic factors,the content of paeoniflorin is positively correlated with latitude and longitude and sunshine hours,and negatively correlated with annual average precipitation and altitude.The areas of southern Xilinguole League of Inner Mongolia Autonomous Region,central and southern Chifeng City and southeast of Hulunbeier and other regions have long sunshine hours,large temperature difference between day and night,and low rainfall is suitable for the growth of Paeoniae Radix Rubra,so it can be used as a suitable planting base for Paeoniae Radix Rubra alternative points.更多还原