SGER: A Method to Genomic Sequence Uncultured Marine Phage

SGER：一种对未培养的海洋噬菌体进行基因组测序的方法

• 批准号: 0116900
• 负责人: Farooq Azam
• 金额: $ 7万
• 依托单位: University of California-San Diego Scripps Inst of Oceanography
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0116900&HistoricalAwards=false
• 关键词: SGER; Method; Genomic; Sequence; Uncultured

Our understanding of the marine environment was fundamentally changed when it was shown that marine phage (viruses that infect bacteria) are present in extraordinary numbers (ca.1010 per liter). Since this observation, marine microbial ecologists have attempted to incorporate phage into the microbial food web concepts. This work has shown that under various conditions phage account for 12-600% of all bacterial production via lysis. The high number of observable phage and their high production rates have called into question concepts of energy and nutrient cycles in the world's oceans. The observation of large numbers of phage in the ocean also raised two other extremely exciting questions: 1) What type of phage inhabit the marine environment?, and 2) How might phage affect genetic transfer (e.g., transduction or lysogenic conversion) in the marine environment?. Available evidence suggests that marine phage diversity is ten times as great as prokaryotic diversity and like their prokaryotic counterparts, it is believed that each phage will have a unique strategy for surviving and multiplying in the marine environment and these strategies will be reflected in the genes that each phage encodes. The life strategies adopted by phage will in turn help shape the MMFW and the biogeochemical processes of the ocean.Current methods of phage characterization rely on phage morphology, genome size, and host range. However, these approaches are extremely limited in their applicability to environmental samples, primarily because the majority of host are not culturable. Drs. Azam and Rohwer ill attempt to circumvent this problem by sequencing the genomes of uncultured phage. This will be accomplished in three steps. First, phage genomes will be separated from each other using pulse field gels. Second, randomly amplified shotgun libraries (RASLs) will be constructed from the bands isolated from the PF-gels. Finally, the shotgun libraries will be sequenced.

当海洋噬菌体（感染细菌的病毒）以惊人的数量（每升约1010个）存在时，我们对海洋环境的理解发生了根本性的变化。自从这一观察以来，海洋微生物生态学家试图将噬菌体纳入微生物食物网的概念。这项工作表明，在各种条件下，噬菌体占所有细菌通过裂解产生的12-600%。大量可观察到的噬菌体及其高生产率使人们对世界海洋中的能量和营养循环概念产生了疑问。对海洋中大量噬菌体的观察还提出了另外两个非常令人兴奋的问题：1）什么类型的噬菌体栖息在海洋环境中？和2）噬菌体如何影响遗传转移（例如，转导或溶原性转化）在海洋环境中？现有的证据表明，海洋噬菌体的多样性是原核生物多样性的十倍，并且像它们的原核生物对应物一样，相信每个噬菌体将具有在海洋环境中生存和繁殖的独特策略，并且这些策略将反映在每个噬菌体编码的基因中。噬菌体所采取的生命策略反过来又会帮助塑造MMFW和海洋的生物地球化学过程。目前的噬菌体表征方法依赖于噬菌体形态、基因组大小和宿主范围。然而，这些方法在其对环境样品的适用性方面极其有限，主要是因为大多数宿主是不可培养的。阿扎姆博士和罗威尔博士将试图通过对未培养的噬菌体的基因组进行测序来规避这个问题。这将分三个步骤完成。首先，使用脉冲场凝胶将噬菌体基因组彼此分离。其次，随机扩增鸟枪文库（RASL）将构建从PF-凝胶分离的条带。最后，将对鸟枪文库进行测序。