Generation and Validation of a Shewanella oneidensis MR-1 Clone Set for Protein Expression and Phage Display

Gao, Haichun; Pattison, Donna; Yan, Tingfen; Klingeman, Dawn M.; Wang, Xiaohu; Petrosino, Joseph; Hemphill, Lisa; Wan, Xiufeng; Leaphart, Adam B.; Weinstock, George M.; Palzkill, Timothy; Zhou, Jizhong

doi:10.1371/journal.pone.0002983

Generation and Validation of a Shewanella oneidensis MR-1 Clone Set for Protein Expression and Phage Display

Journal Article · Wed Aug 20 00:00:00 EDT 2008 · PLoS ONE

DOI:https://doi.org/10.1371/journal.pone.0002983· OSTI ID:1627357

Gao, Haichun ^[1]; Pattison, Donna ^[2]; Yan, Tingfen ^[3]; Klingeman, Dawn M. ^[3]; Wang, Xiaohu ^[2]; Petrosino, Joseph ^[2]; Hemphill, Lisa ^[2]; Wan, Xiufeng ^[3]; Leaphart, Adam B. ^[3]; Weinstock, George M. ^[2]; Palzkill, Timothy ^[2]; Zhou, Jizhong ^[4]

Univ. of Oklahoma, Norman, OK (United States). Inst. for Environmental Genomics; Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Environmental Sciences Division; DOE/OSTI
Baylor Univ., Houston, TX (United States). College of Medicine
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Environmental Sciences Division
Univ. of Oklahoma, Norman, OK (United States). Inst. for Environmental Genomics; Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Environmental Sciences Division

A comprehensive gene collection for S. oneidensis was constructed using the lambda recombinase (Gateway) cloning system. A total of 3584 individual ORFs (85%) have been successfully cloned into the entry plasmids. To validate the use of the clone set, three sets of ORFs were examined within three different destination vectors constructed in this study. Success rates for heterologous protein expression of S. oneidensis His- or His/GST- tagged proteins in E. coli were approximately 70%. The ArcA and NarP transcription factor proteins were tested in an in vitro binding assay to demonstrate that functional proteins can be successfully produced using the clone set. Further functional validation of the clone set was obtained from phage display experiments in which a phage encoding thioredoxin was successfully isolated from a pool of 80 different clones after three rounds of biopanning using immobilized anti-thioredoxin antibody as a target. This clone set complements existing genomic (e.g., whole-genome microarray) and other proteomic tools (e.g., mass spectrometry-based proteomic analysis), and facilitates a wide variety of integrated studies, including protein expression, purification, and functional analyses of proteins both in vivo and in vitro.

View Accepted Manuscript (DOE)

Research Organization:: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division

Grant/Contract Number:: AC05-00OR22725

OSTI ID:: 1627357

Journal Information:: PLoS ONE, Journal Name: PLoS ONE Journal Issue: 8 Vol. 3; ISSN 1932-6203

Publisher:: Public Library of ScienceCopyright Statement

Country of Publication:: United States

Language:: English

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Transcriptome Analysis Applied to Survival of Shewanella oneidensis MR-1 Exposed to Ionizing Radiation Qiu, X.; Daly, M. J.; Vasilenko, A. Journal of Bacteriology, Vol. 188, Issue 3 https://doi.org/10.1128/JB.188.3.1199-1204.2006	journal	January 2006
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Impaired cell envelope resulting from arcA mutation largely accounts for enhanced sensitivity to hydrogen peroxide in Shewanella oneidensis Wan, Fen; Mao, Yinting; Dong, Yangyang Scientific Reports, Vol. 5, Issue 1 https://doi.org/10.1038/srep10228	journal	May 2015
Physiological Roles of ArcA, Crp, and EtrA and Their Interactive Control on Aerobic and Anaerobic Respiration in Shewanella oneidensis Gao, Haichun; Wang, Xiaohu; Yang, Zamin K. PLoS ONE, Vol. 5, Issue 12 https://doi.org/10.1371/journal.pone.0015295	journal	December 2010
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Transcription factors FabR and FadR regulate both aerobic and anaerobic pathways for unsaturated fatty acid biosynthesis in Shewanella oneidensis Luo, Qixia; Shi, Miaomiao; Ren, Yedan Frontiers in Microbiology, Vol. 5 https://doi.org/10.3389/fmicb.2014.00736	journal	December 2014
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