箭竹属的 DNA 条形码研究

	箭竹属的 DNA 条形码研究
	吕时雨
导师	李德铢
关键词	箭竹属，浅层测序， DNA条形码，叶绿体基因组，核糖体DNA Fargesia, genome-skimming, DNA barcoding, plastome, ribosomal DNA
摘要	箭竹属（Fargesia）隶属于禾本科竹亚科（Poaceae: Bambusoideae）青篱竹族（Arundinarieae），是温带木本竹中最大的属，该属包含90余种，主要分布于中国西南部山区。箭竹属植物是森林生态系统的重要组成部分，也是一些濒危野生动物如大熊猫等的重要食物来源，具有重要的生态及经济价值。然而，由于竹类植物的开花周期长、繁殖器官难以获得以及营养器官性状不稳定等因素，导致形态学的物种鉴定十分困难。此外，标准植物DNA条形码（matK, rbcL, trnH-psbA）在该类植物中鉴定成功率极低，难以达到分子鉴定的目的。随着测序技术的发展和条形码研究的不断深入，全叶绿体基因组和核糖体DNA序列已被提议作为第二代植物DNA条形码（plant DNA barcode 2.0）用于近缘物种的鉴定，也被称为超级条形码（ultra-barcode），但尚未应用于竹类植物中。本研究共收集了62个箭竹属物种共196个样品，通过与标准条形码的比较来全面探讨全叶绿体基因组和核糖体DNA序列在箭竹属物种鉴定中的应用。本研究新获得了129个完整的箭竹属叶绿体基因组以及110条核糖体DNA序列，并从GenBank下载了7个叶绿体基因组和22条ITS序列作为补充，共构建了七个数据集，每个数据集使用基于系统进化树和基于遗传距离的两种方法来计算物种鉴定成功率。在叶绿体DNA条形码中，叶绿体标准条形码的组合序列matK + rbcL + trnH-psbA的鉴定率最低，仅能鉴定5.7%的物种，全叶绿体基因组以及LSC区域的鉴定率最高，达到了28.6%。与叶绿体标准条形码相比，全叶绿体基因组可以显著的提高箭竹属的鉴定效率，但叶绿体基因组序列的变异率极低，即便使用全叶绿体基因组序列，物种的成功鉴定率仍然很低。而基于双亲遗传的核基因ITS及核糖体DNA序列能得到更高的鉴定成功率，仅ITS序列的鉴定率就远高于全叶绿体基因组序列，达到了47.2%。核糖体DNA的鉴定率相比于ITS也有适度的提高，鉴定率为57.7%。研究结果表明，在箭竹属中，基于新一代测序技术获得的全叶绿体基因组和核糖体DNA序列能够显著的提高物种鉴定成功率。但是，即使使用鉴定率最高的核糖体DNA序列，箭竹属中仍有很多物种不能成功鉴定，造成箭竹属物种鉴定率低的主要原因可能是该类群漫长的世代时间以及复杂的演化历史。因此，本研究再次强化了核基因组数据在物种鉴定中的重要性。; Fargesia, the largest genus within the temperate bamboos tribe Arundinarieae (Poaceae: Bambusoideae), has more than 90 species, mainly distributed in the mountains of Southwest China. These bamboos have considerable ecological and economic value, they are essential components of the alpine forest ecosystem, which provides food and habitat for many endangered animals, including the giant panda. However, due to the long flowering intervals and variable morphological characteristics, it is peculiarly difficult to identify Fargesia species depending on morphology alone. Moreover, the discriminatory power of standard DNA barcodes (matK, rbcL, trnH-psbA) in identifying bamboos was very low, it is far from adequate for achieving DNA identification in bamboos. With the development of new sequencing approaches, complete plastid genome (plastome) and ribosomal DNA (nrDNA) have been proposed as candidates for the next generation of DNA barcodes (plant DNA barcode 2.0) to increase the power of plant species discrimination, also known as ultra-barcode. However, ultra-barcode has not been tested in bamboos. In this study, we collected 196 individuals representing 62 species of Fargesia to comprehensively evaluate the discriminatory power of plastome and nrDNA sequences compared to the standard barcodes. We newly obtained 136 plastomes and 110 nrDNA sequences for Fargesia. Another, 7 individuals with whole plastomes sequences and 22 individuals with ITS sequences were downloaded from the GenBank. We constructed seven datasets to compare the discriminatory power. To evaluate species discrimination success, we used two approaches: tree-based methods and distance-based analysis. For DNA barcodes based on plastid data, Three-marker combinations of the plastid barcodes (matK+rbcL+trnH-psbA) showed the lowest discriminatory power in Fargesia with the discrimination rate at 5.7%, the complete plastomes and LSC region showed the highest discrimination rate at 28.6%. Though the complete plastome can substantially increase discriminatory power compared to standard plastid barcodes, the slow substitution rates of Fargesia plastomes result in the discrimination rate remaining low. We also found nuclear markers performed better than plastid markers, and even ITS alone had higher discriminatory power (47.2%) than complete plastome. It showed a moderate improvement when comparing nrDNA to ITS, and nrDNA sequences show a moderate improvement compared to ITS with 57.7% of species distinguished. Our analysis found that the complete plastome and nrDNA sequences had substantially higher discriminatory power than the standard barcodes in Fargesia. However, neither of these sequences could discriminate all the sampled species, possibly due to the long generation time and complex evolutionary history of Fargesia. Therefore, our results reinforce the need to access multiple nuclear markers in species discrimination in plants.
语种	中文
	2022-06
学位授予单位	中国科学院大学
文献类型	学位论文
条目标识符	http://ir.kib.ac.cn/handle/151853/75172
专题	昆明植物所硕博研究生毕业学位论文
推荐引用方式 GB/T 7714	吕时雨. 箭竹属的 DNA 条形码研究[D]. 中国科学院大学,2022.

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