Global Transposon Mutagenesis and a Minimal Mycoplasma Genome

Hutchison, Clyde A.; Peterson, Scott N.; Gill, Steven R.; Cline, Robin T.; White, Owen; Fraser, Claire M.; Smith, Hamilton O.; Venter, J. Craig

doi:10.1126/science.286.5447.2165

Cited by 825 publications

(619 citation statements)

References 27 publications

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“…This gene has been shown to be nonessential in M. genitalium, but not in M. pneumoniae. However, the vector insertion occurred near the 3 0 -end of the gene, which may not be disruptive to gene function (44,45). The restriction map produced by vector insertion at this location matched our results.…”

Section: Cloning Mycoplasma Genomes That Contain An Integrated Yeast supporting

confidence: 84%

Cloning Whole Bacterial Genomes in Yeast

Benders

2012

Methods in Molecular Biology

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Most microbes have not been cultured, and many of those that are cultivatable are difficult, dangerous or expensive to propagate or are genetically intractable. Routine cloning of large genome fractions or whole genomes from these organisms would significantly enhance their discovery and genetic and functional characterization. Here we report the cloning of whole bacterial genomes in the yeast Saccharomyces cerevisiae as single-DNA molecules. We cloned the genomes of Mycoplasma genitalium (0.6 Mb), M. pneumoniae (0.8 Mb) and M. mycoides subspecies capri (1.1 Mb) as yeast circular centromeric plasmids. These genomes appear to be stably maintained in a host that has efficient, well-established methods for DNA manipulation.

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Section: Cloning Mycoplasma Genomes That Contain An Integrated Yeast supporting

confidence: 84%

Cloning Whole Bacterial Genomes in Yeast

Benders

2012

Methods in Molecular Biology

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“…Interestingly, we found that in the pilot phase of PSI, the families that were first structurally characterized by the BSGC averaged twice the size of the families characterized by the other 8 pilot centers, containing an average of 262 members, vs. 130 members for the other centers [29]. This is presumably a result of focusing on a minimal organism, as a large fraction of M. genitalium's genes are thought to be essential for life, and therefore nearly ubiquitous across a wide range of species [36].…”

Section: Novelty Of Bsgc Structuresmentioning

confidence: 99%

Structure-based inference of molecular functions of proteins of unknown function from Berkeley Structural Genomics Center

Shin

Hou

Chandonia

et al. 2007

J Struct Funct Genomics

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As a part of the Protein Structure Initiative (PSI; www.nigms.nih.gov/funding/psi.html) the BSGC has focused on obtaining the 3D structural information of the proteins of two minimal organisms, closely related pathogens Mycoplasma genitalium and M. pneumoniae (http://www.strgen.org), which have fewer than 500 and 700 genes, respectively. The requisite to achieve this goal involved obtaining 3D structural information for nearly all proteins, a large portion of which are hypothetical proteins, the proteins with no sequence homologies to those of known function. Now, we have 3D structural information for near complete structural complement of M. genitalium. Thus, we now have a structural genomic view of protein fold usage among these and other minimal microbes [3]. 3 Metrics and Impact of BSGC structuresAt the beginning of PSI initiative, about 30% of M. genitalium "soluble" proteins had no 3-D structural fold information. By the time of the completion of the PSI-I, about 94% of the "soluble" proteins have 3-D structural fold information, thus, achieving the mission of BSGC of obtaining a near complete structural complement of a minimal organism. Several metrics were learned from the exercise:(1) About 1/2 of proteins that had no sequence similarity to the proteins in PDB turned out to have "new folds" and ~1/2 turned out to be "remote homologues" in which homology could only be identified through structural similarity to a known fold.(2) About 2/3 of the 3-D structures of "hypothetical" proteins inferred testable molecular (biochemical or biophysical) functions, and some of which have since been confirmed experimentally.(3) The overall success rate of "single-path" (low-hanging fruit) approach for clone-to-structure was <5%, and for purified protein-to-structure was ~9%.(4) The overall success rate of "multi-path" (single-path plus "salvage path") approach for clone-to-structure was >16%, and for purified protein-to-structure was ~27% 4The overall impact of BSGC structures to the functional inference is summarized below:• 66 BSGC structures belong to 51 protein sequence families.

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“…When Hutchison took a sabbatical at TIGR in 1996, he, Smith and Venter began to discuss the idea of developing a cell with the minimum genome needed to survive. Hutchison was already investigating which genes M. genitalium could live without 6 , but he knew that deleting multiple genes simultaneously from this bacterium was technically difficult. The threesome speculated that they might need to synthesize whole candidate genomes and test them in recipient cells.…”

Section: Behind the Walls Of The J Craig Venter Institute Ham Smithmentioning

confidence: 99%

Genomics: DNA's master craftsmen

Kwok¹

2010

Nature

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Global Transposon Mutagenesis and a Minimal Mycoplasma Genome

Cited by 825 publications

References 27 publications

Cloning Whole Bacterial Genomes in Yeast

Cloning Whole Bacterial Genomes in Yeast

Structure-based inference of molecular functions of proteins of unknown function from Berkeley Structural Genomics Center

Genomics: DNA's master craftsmen

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