J P Tomkins, H Miller-Smith, M Sasinowski, S Choi, H Sasinowska, M F Verce, D L Freedman, R A Dean, R A Wing
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In addition, 1536 BAC clones were fingerprinted with HindIII and analyzed using IMAGE and Fingerprint Contig software (FPC, Sanger Centre, U.K.). The FPC results supported the hybridization data, resulting in the formation of two major contigs representing the two major replicons of the O. anthropi genome. After determining a reduced tiling path, 138 BAC ends from the reduced tile were sequenced for a preliminary gene survey. A search of the public databases with the BLASTX algorithm resulted in 77 strong hits (E-value < 0.001), of which 89% showed similarity to a wide variety of prokaryotic genes. 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引用次数: 8
摘要
细菌人工染色体(BAC)克隆是一种有效的定位和测序试剂,可用于各种大小基因组。本报告描述了旨在确定人类赭杆菌基因组结构的研究,这是一种机会性人类病原体,在有害有机化合物的生物降解中具有潜在的应用。基于估算的基因组大小为4.8 Mb,构建了人类猿猴BAC文库,该文库包含3072个克隆,平均插入长度为112 kb。制作该文库的高密度菌落过滤器,并采用无置换杂交策略构建基因组物理图谱。此外,对1536个BAC克隆进行HindIII指纹鉴定,并使用IMAGE和Fingerprint Contig软件(FPC, Sanger Centre, U.K.)进行分析。FPC结果支持杂交数据,结果形成了两个主要的contigs,代表了猿人基因组的两个主要复制子。在确定还原瓦片路径后,对来自还原瓦片的138个BAC端进行了初步的基因调查。使用BLASTX算法对公共数据库进行搜索,结果得到77个强命中(e值< 0.001),其中89%显示与多种原核基因相似。这些结果为类人猿重要基因组区域的克隆和潜在的基因组测序提供了一个基于组群的物理图谱。
Physical map and gene survey of the Ochrobactrum anthropi genome using bacterial artificial chromosome contigs.
Bacterial artificial chromosome (BAC) clones are effective mapping and sequencing reagents for use with a wide variety of small and large genomes. This report describes research aimed at determining the genome structure of Ochrobactrum anthropi, an opportunistic human pathogen that has potential applications in biodegradation of hazardous organic compounds. A BAC library for O. anthropi was constructed that provides a 70-fold genome coverage based on an estimated genome size of 4.8 Mb. The library contains 3072 clones with an average insert size of 112 kb. High-density colony filters of the library were made, and a physical map of the genome was constructed using a hybridization without replacement strategy. In addition, 1536 BAC clones were fingerprinted with HindIII and analyzed using IMAGE and Fingerprint Contig software (FPC, Sanger Centre, U.K.). The FPC results supported the hybridization data, resulting in the formation of two major contigs representing the two major replicons of the O. anthropi genome. After determining a reduced tiling path, 138 BAC ends from the reduced tile were sequenced for a preliminary gene survey. A search of the public databases with the BLASTX algorithm resulted in 77 strong hits (E-value < 0.001), of which 89% showed similarity to a wide variety of prokaryotic genes. These results provide a contig-based physical map to assist the cloning of important genomic regions and the potential sequencing of the O. anthropi genome.