布渣叶种内遗传分化与活性成份变异的地理格局及其形成机制

It is well known that the active metabolic products of medical plant are the effective material basis for their medical uses, and their variations are jointly affected by genetic and environmental factors. An in depth knowledge of the inner correlationship between genetic and chemical diversity and environmental parameters can help us better understand the quality formation mechanism of the raw herbs, and then screen and breed excellent germplasm for GAP cultivation. Based on the discover in the past research that rich chemical diversity lies among Microcos paniculata L. populations, we intend to analysis their genetic structure and genetic diversity of 16 populations scattering over Guangdong Province, Guangxi Province and Hainan Province by ISSR molecular marker technology, and establish a dataset which was composed of all the genetic diversity parameters calculated by computer software. At the same time, the acitve metabolic substance and their variation in corresponding samples collected from the 16 Microcos paniculata L. populations were analyzed by UPLC-MS/MS and HPLC, and as well, a dataset which characterized the chemical diversity was established. Then, the associations between the two dataset and the geographical and climatological factors were evaluated by principal component analysis, discriminant analysis, partial least-squares regression and other multivariate statistical analysis. So, the results of our research can reveal the intraspecies genetic differentiation and spatio-chemical variation pattern and their mechanism of Microcos paniculata L. on population lever, and set a theoretical foundation for germplasm resource evaluation, core germplasm protection as well as breeding.

药用植物的活性代谢产物是其发挥药效的基础，其多样性变化受遗传物质和生境因子的共同作用，充分研究物种的遗传多样性、活性代谢物质的化学多样性与生境因子的内在联系，有助于理解药材品质形成机制，进而选育优良种质。本项目在发现布渣叶代谢产物存在丰富的化学多样性基础上，拟通过ISSR分子标记分析广东、广西、海南等16个布渣叶居群遗传结构/遗传多样性，建立反映其遗传多样性的数据集；同时，运用 UPLC-MS/MS和HPLC分析上述样品的活性代谢物质及其变化，建立表征其化学多样性的数据集；然后将二者与地理气候因子通过主成份分析、判别分析、偏最小二乘回归等多元统计方法关联起来，试图通过这些研究来阐明布渣叶居群遗传结构/遗传多样性、活性代谢产物变异与地理气候因子的相互关系，从而揭示布渣叶种内遗传分化和化学变异地理格局以及相应的基因流机制和生境影响机制，为其种质资源评价，核心种质保护和良种的选育提供理论依据。

布渣叶，岭南特色“药食两用”植物，广东凉茶饮料和保儿安颗粒、胃肠宁颗粒等中成药的生产原料，市场需求量大而鲜有栽培，资源破坏日趋严重，令人堪忧。在项目研究过程中，课题组取得了如下进展及研究结果：（1）建立了布渣叶DNA条形码技术体系，为其基源植物和药材饮片的分子鉴定提供了技术支撑；（2）从布渣叶中分离、纯化和鉴定了20个黄酮及酚酸类化合物，其中首次从该属植物中得到的化合物6个，为其药效物质基础研究提供了参考；（3）对于布渣叶及其提取物，建立了基于HPLC的指纹图谱以及“一测多评”方法，同时测定其中的8种黄酮苷类化合物，为其质量评价体系建立提供了依据；（4）对不同种源布渣叶主要活性成分变化与其生境和遗传因素的相关性分析发现，日照时间对布渣叶黄酮苷类积累量影响十分显著，与部分化合物的含量变化呈明显相关性，为其品质形成的生态学影响提供了理论依据；（5）从广东、广西、海南和云南等地14个自然居群的布渣叶样本中筛选出较好的布渣叶种源地2个和优良种质2份，为其良种选育和核心种质构建奠定了基础；（6）对布渣叶优良种质的转录组de novo组装和分析，发现了多个候选的布渣叶黄酮类生物合成相关的功能基因、转录因子；并对 Mp4CL，MpCHS，MpF2H等基因进行了克隆、生物信息学分析和原核表达。上述研究结果，不但解决了布渣叶基源植物及其药材饮片的分子鉴定技术问题，而且利用HPLC、LC-MS等现代分析方法，建立了其质量评价体系；对布渣叶的资源评价和种质评鉴具有重要的参考意义和应用价值。同时，初步阐明了布渣叶活性成分变异与其遗传及生境因素的相关性，亦筛选出相应的次生代谢途径上的关键酶基因/转录因子等，对揭示生境及遗传对布渣叶次生代谢的影响，阐释其品质形成的分子机制具有重要的科学意义，为后续布渣叶种质创新研究奠定了坚实的基础，对其他药用植物芹黄素-C-苷类成分的生物合成研究也具有重要的参考价值。