BMC Genomics 2009, 10:165 http://www.biomedcentral.com/1471-2164/10/165 data analysis and editing of the manuscript. All authors read and approved the final manuscript. Additional material Additional file 1 Small noncoding RNAs (sncRNA) that were selected for construction of a single-sncRNA gene knockout library within E. coli K12- MG1655. This table summarizes positions and functions of the small non- coding RNAs that were included in a single-sncRNA gene knockout library within E. coli K12-MG1655. Click here for file [http://www.biomedcentral.com/content/supplementary/1471- 2164-10-165-S1.doc] Acknowledgements We thank D.L. Court for providing plasmid pSim6 for the recombineering experiments. This work was supported by HKU CRCG Seed Funding Pro- gramme for Basic Research and by a grant (HKU 7485/06M) from the Research Grant Council of Hong Kong to JDH. AD acknowledges support from the PROBACTYS programme, grant CT-2006-029104 in an effort to define genes essential for the construction of a synthetic cell. References 1. Castanie-Cornet MP, Penfound TA, Smith D, Elliott JF, Foster JW: Control of acid resistance in Escherichia coli. J Bacteriol 1999, 181(11):3525-3535. 2. Hersh BM, Farooq FT, Barstad DN, Blankenhorn DL, Slonczewski JL: A glutamate-dependent acid resistance gene in Escherichia coli. J Bacteriol 1996, 178(13):3978-3981. 3. Lin J, Smith MP, Chapin KC, Baik HS, Bennett GN, Foster JW: Mech- anisms of acid resistance in enterohemorrhagic Escherichia coli. Appl Environ Microbiol 1996, 62(9):3094-3100. 4. Sledjeski DD, Gupta A, Gottesman S: The small RNA, DsrA, is essential for the low temperature expression of RpoS during exponential growth in Escherichia coli. EMBO J 1996, 15(15):3993-4000. 5. Lease RA, Smith D, McDonough K, Belfort M: The small noncod- ing DsrA RNA is an acid resistance regulator in Escherichia coli. J Bacteriol 2004, 186(18):6179-6185. 6. Opdyke JA, Kang JG, Storz G: GadY, a small-RNA regulator of acid response genes in Escherichia coli. J Bacteriol 2004, 186(20):6698-6705. 7. Tramonti A, De Canio M, De Biase D: GadX/GadW-dependent regulation of the Escherichia coli acid fitness island: tran- scriptional control at the gadY-gadW divergent promoters and identification of four novel 42-bp GadX/GadW-specific binding sites. Mol Microbiol 2008, 70(4):965-982. 8. Griffiths-Jones S, Moxon S, Marshall M, Khanna A, Eddy SR, Bateman A: Rfam: annotating non-coding RNAs in complete genomes. Nucleic Acids Res 2005:D121-124. 9. Vallenet D, Labarre L, Rouy Z, Barbe V, Bocs S, Cruveiller S, Lajus A, Pascal G, Scarpelli C, Medigue C: MaGe: a microbial genome annotation system supported by synteny results. Nucleic Acids Res 2006, 34(1):53-65. 10. Vogel J, Bartels V, Tang TH, Churakov G, Slagter-Jager JG, Hutten- hofer A, Wagner EG: RNomics in Escherichia coli detects new sRNA species and indicates parallel transcriptional output in bacteria. Nucleic Acids Res 2003, 31(22):6435-6443. 11. Ellis HM, Yu D, DiTizio T, Court DL: High efficiency mutagenesis, repair, and engineering of chromosomal DNA using single- stranded oligonucleotides. Proc Natl Acad Sci USA 2001, 98(12):6742-6746. 12. Yu D, Ellis HM, Lee EC, Jenkins NA, Copeland NG, Court DL: An efficient recombination system for chromosome engineer- ing in Escherichia coli. Proc Natl Acad Sci USA 2000, 97(11):5978-5983. 13. Urbanowski ML, Stauffer LT, Stauffer GV: The gcvB gene encodes a small untranslated RNA involved in expression of the dipeptide and oligopeptide transport systems in Escherichia coli. Mol Microbiol 2000, 37(4):856-868. 14. Urban JH, Vogel J: Translational control and target recognition by Escherichia coli small RNAs in vivo. Nucleic Acids Res 2007, 35(3):1018-1037. 15. Sharma CM, Darfeuille F, Plantinga TH, Vogel J: A small RNA reg- ulates multiple ABC transporter mRNAs by targeting C/A- rich elements inside and upstream of ribosome-binding sites. Genes Dev 2007, 21(21):2804-2817. 16. Tramonti A, Visca P, De Canio M, Falconi M, De Biase D: Functional characterization and regulation of gadX, a gene encoding an AraC/XylS-like transcriptional activator of the Escherichia coli glutamic acid decarboxylase system. J Bacteriol 2002, 184(10):2603-2613. 17. Sayed AK, Odom C, Foster JW: The Escherichia coli AraC-fam- ily regulators GadX and GadW activate gadE, the central activator of glutamate-dependent acid resistance. Microbiol- ogy 2007, 153(Pt 8):2584-2592. 18. Small P, Blankenhorn D, Welty D, Zinser E, Slonczewski JL: Acid and base resistance in Escherichia coli and Shigella flexneri: role of rpoS and growth pH. J Bacteriol 1994, 176(6):1729-1737. 19. Mika F, Hengge R: A two-component phosphotransfer network involving ArcB, ArcA, and RssB coordinates synthesis and proteolysis of sigmaS (RpoS) in E. coli. Genes Dev 2005, 19(22):2770-2781. 20. Lange R, Hengge-Aronis R: The cellular concentration of the sigma S subunit of RNA polymerase in Escherichia coli is controlled at the levels of transcription, translation, and pro- tein stability. Genes Dev 1994, 8(13):1600-1612. 21. Mukhopadhyay S, Audia JP, Roy RN, Schellhorn HE: Transcriptional induction of the conserved alternative sigma factor RpoS in Escherichia coli is dependent on BarA, a probable two-com- ponent regulator. Mol Microbiol 2000, 37(2):371-381. 22. Hommais F, Krin E, Coppee JY, Lacroix C, Yeramian E, Danchin A, Bertin P: GadE (YhiE): a novel activator involved in the response to acid environment in Escherichia coli. Microbiology 2004, 150(Pt 1):61-72. 23. Brescia CC, Kaw MK, Sledjeski DD: The DNA binding protein H- NS binds to and alters the stability of RNA in vitro and in vivo. J Mol Biol 2004, 339(3):505-514. 24. Lease RA, Cusick ME, Belfort M: Riboregulation in Escherichia coli: DsrA RNA acts by RNA:RNA interactions at multiple loci. Proc Natl Acad Sci USA 1998, 95(21):12456-12461. 25. Zhang A, Altuvia S, Tiwari A, Argaman L, Hengge-Aronis R, Storz G: The OxyS regulatory RNA represses rpoS translation and binds the Hfq (HF-I) protein. EMBO J 1998, 17(20):6061-6068. 26. Lange R, Fischer D, Hengge-Aronis R: Identification of transcrip- tional start sites and the role of ppGpp in the expression of rpoS, the structural gene for the sigma S subunit of RNA polymerase in Escherichia coli. J Bacteriol 1995, 177(16):4676-4680. 27. Majdalani N, Cunning C, Sledjeski D, Elliott T, Gottesman S: DsrA RNA regulates translation of RpoS message by an anti-anti- sense mechanism, independent of its action as an antisi- lencer of transcription. Proc Natl Acad Sci USA 1998, 95(21):12462-12467. 28. Majdalani N, Hernandez D, Gottesman S: Regulation and mode of action of the second small RNA activator of RpoS transla- tion, RprA. Mol Microbiol 2002, 46(3):813-826. 29. Updegrove T, Wilf N, Sun X, Wartell RM: Effect of Hfq on RprA- rpoS mRNA pairing: Hfq-RNA binding and the influence of the 5' rpoS mRNA leader region. Biochemistry 2008, 47(43):11184-11195. 30. Resch A, Afonyushkin T, Lombo TB, McDowall KJ, Blasi U, Kaberdin VR: Translational activation by the noncoding RNA DsrA involves alternative RNase III processing in the rpoS 5'- leader. RNA 2008, 14(3):454-459. 31. Kruger J, Rehmsmeier M: RNAhybrid: microRNA target predic- tion easy, fast and flexible. Nucleic Acids Res 2006:W451-454. 32. Datta S, Costantino N, Court DL: A set of recombineering plas- mids for gram-negative bacteria. Gene 2006, 379:109-115. Page 7 of 7 (page number not for citation purposes)