Metatranscriptomic analysis of autonomously collected and preserved marine bacterioplankton
Metatranscriptomic analysis of autonomously collected and preserved marine bacterioplankton
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DOI：
10.1038/ismej.2011.70
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发表时间：
2011-12-01
期刊：
影响因子：
11
通讯作者：
DeLong, Edward F.
中科院分区：
文献类型：
--
作者：
Ottesen, Elizabeth A.;Marin, Roman, III;DeLong, Edward F.
浮游微生物的活动和群落结构是动态的，可以在几小时到几天的时间尺度上发生巨大变化。然而，出于后勤原因，在海洋环境中，这种时间尺度通常是采样不足的。为了便于对微生物多样性和活性进行更高分辨率的长期观察，我们开发了一种使用环境样品处理器(ESP)平台自动收集和固定海洋微生物的协议。该协议将一种防腐剂(RNAlater)应用于通过过滤器收集的细胞，以长期存储和保存细胞总RNA。用这种方法保存的微生物样本在室温下储存30天后产生高质量的RNA，或者在原位温度下安装在ESP上。对从ESP收集和保存的样本产生的互补DNA文库进行焦磷酸测序，得到的转录丰度谱与那些来自常规处理的复制样本的转录本丰度谱几乎没有区别。为了证明这种方法的实用性，我们使用系泊的电除尘器远程和自主地收集蒙特利湾的海水进行元转录分析。从一天中的四个时间点收集的样本中提取社区RNA并进行焦解测序。在所有4个样品中，产氧光自养细菌以真核生物为主，细菌群落以类Polaribacter黄杆菌和一株与红杆菌属有很高相似性的红杆菌属细菌为主。HTCC2255。然而，每个时间点都与不同的物种丰度和基因转录谱相关。这些实验室和野外测试证实，自主收集和保存是表征海洋微生物群落表达的基因和环境响应的一种可行和有用的方法。《ISME期刊》(2011年)第5期，1881年至1895年；doi：10.1038/ismej.2011.70；2011年6月30日在线出版
Planktonic microbial activity and community structure is dynamic, and can change dramatically on time scales of hours to days. Yet for logistical reasons, this temporal scale is typically under-sampled in the marine environment. In order to facilitate higher-resolution, long-term observation of microbial diversity and activity, we developed a protocol for automated collection and fixation of marine microbes using the Environmental Sample Processor (ESP) platform. The protocol applies a preservative (RNALater) to cells collected on filters, for long-term storage and preservation of total cellular RNA. Microbial samples preserved using this protocol yielded high-quality RNA after 30 days of storage at room temperature, or onboard the ESP at in situ temperatures. Pyrosequencing of complementary DNA libraries generated from ESP-collected and preserved samples yielded transcript abundance profiles nearly indistinguishable from those derived from conventionally treated replicate samples. To demonstrate the utility of the method, we used a moored ESP to remotely and autonomously collect Monterey Bay seawater for metatranscriptomic analysis. Community RNA was extracted and pyrosequenced from samples collected at four time points over the course of a single day. In all four samples, the oxygenic photoautotrophs were predominantly eukaryotic, while the bacterial community was dominated by Polaribacter-like Flavobacteria and a Rhodobacterales bacterium sharing high similarity with Rhodobacterales sp. HTCC2255. However, each time point was associated with distinct species abundance and gene transcript profiles. These laboratory and field tests confirmed that autonomous collection and preservation is a feasible and useful approach for characterizing the expressed genes and environmental responses of marine microbial communities. The ISME Journal (2011) 5, 1881-1895; doi: 10.1038/ismej.2011.70; published online 30 June 2011