[Two sample pooling strategies revealed different root-associated fungal diversity of Rhododendron species].

微生物学报 Pub Date : 2017-04-04
Caiwei Huang, Yinghui Liao, Qiong Ding
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引用次数: 0

Abstract

Objective: ooling of multiple samples is widely used in studying general patterns of microbial communities that are heterogeneously structured in space. Pooling strategies and the number of sequence reads generate biases in the description of diversity and community structure of root-associated fungi. Therefore, we developed a molecular toolbox for fast and accurate identification of the root-associated fungal community of Rododendron species.

Methods: Multiple root samples of R. lutescens and R. bureavii were collected for DNA extraction. Effects of two different pooling strategies, i.e. pooling samples prior to vs. post PCR, on fungal species composition were studied by comparing results within host species.

Results: Species richness and Shannon-Wiener index of fungal communities of clone library constructed by pooling samples after PCR were higher than that of pooling prior to PCR. High frequency fungal species were detected by both pooling strategies, whereas infrequent species detected by the two strategies differed. Notably, the prior to PCR pooling strategy effectively alleviated the unwanted amplification of host plant sequences when fungal specific primer ITS1f and ITS4 were used. Accumulation curves of fungal species suggested that sequencing at least 50 clones can fully reflect species composition of clone library of the two Rhododendron root-associated fungal community.

Conclusion: Clone library constructed by post PCR pooling of samples is better in providing accurate views of fungal diversity and community structure of Rhododendron species.

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[两种样本池策略揭示了不同杜鹃花物种根相关真菌多样性]。
目的:多样品冷却被广泛应用于研究空间异质性结构的微生物群落的一般模式。池化策略和序列读取数在描述根相关真菌的多样性和群落结构时会产生偏差。因此,我们开发了一个分子工具箱,用于快速准确地鉴定杜鹃属植物的根相关真菌群落。方法:采集多根木犀草和布氏木犀草进行DNA提取。通过比较宿主物种内的结果,研究了两种不同的池化策略(即PCR前和PCR后的池化策略)对真菌物种组成的影响。结果:PCR后池化构建的克隆文库真菌群落的物种丰富度和Shannon-Wiener指数均高于PCR前池化构建的克隆文库。两种方法均能检测到高频真菌种类,而两种方法检测到的低频率真菌种类不同。值得注意的是,当使用真菌特异性引物ITS1f和ITS4时,预先PCR池策略有效地减轻了寄主植物序列的不必要扩增。真菌物种积累曲线表明,至少50个克隆的测序可以充分反映两个杜鹃根相关真菌群落克隆文库的物种组成。结论:采用聚合酶链反应(post - PCR)方法构建的克隆文库能够较准确地了解杜鹃属植物的真菌多样性和群落结构。
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期刊介绍: Acta Microbiologica Sinica(AMS) is a peer-reviewed monthly (one volume per year)international journal,founded in 1953.It covers a wide range of topics in the areas of general and applied microbiology.The journal publishes original papers,reviews in microbiological science,and short communications describing unusual observations. Acta Microbiologica Sinica has been indexed in Index Copernicus (IC),Chemical Abstract (CA),Excerpt Medica Database (EMBASE),AJ of Viniti (Russia),Biological Abstracts (BA),Chinese Science Citation Database (CSCD),China National Knowledge Infrastructure(CNKI),Institute of Scientific and Technical Information of China(ISTIC),Chinese Journal Citation Report(CJCR),Chinese Biological Abstracts,Chinese Pharmaceutical Abstracts,Chinese Medical Abstracts and Chinese Science Abstracts.
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