Abstract

This proposal is to finish the sequence of the entire chromosome of the bacterium E. coli. The sequence will be determined using the Sanger dideoxy method implemented on AB fluorescence sequencing machines. The strategy chosen entails an initial phase of random shotgun sequencing followed by reverse strand sequencing by the Janus vector and finally primer walking to achieve closure. A novel method is proposed for isolating suitable fragments of the genome to avoid resequencing already sequenced portions. These fragments will be defined by pairs of unique specific endonuclease cut sites introduced into the genome through mini- transposon metagenesis and will be isolated in high yield through preparative pulse field gel electrophoresis. The backup strategy entails sequencing already available lambda clones of E. coli DNA. The sequence of E. coli, when completed, will provide a major resource for understanding the genetics of the bacterial cell and by extension, of the human cell, for many of the genes of human will find homologs in the bacterial sequence. The sequence will also be of value in understanding the mechanisms of bacterial pathogenesis.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Research Program Projects (P01)
Project #
1P01HG001428-01
Application #
2209751
Study Section
Special Emphasis Panel (SRC (01))
Project Start
1995-09-30
Project End
1997-06-30
Budget Start
1995-09-30
Budget End
1996-06-30
Support Year
1
Fiscal Year
1995
Total Cost
Indirect Cost

  Institution

Name
University of Wisconsin Madison
Department
Other Health Professions
Type
Schools of Earth Sciences/Natur
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715

  Publications

Rotman, Ella; Kouzminova, Elena; Plunkett 3rd, Guy et al. (2012) Genome of Enterobacteriophage Lula/phi80 and insights into its ability to spread in the laboratory environment. J Bacteriol 194:6802-17
Perry, R D; Straley, S C; Fetherston, J D et al. (1998) DNA sequencing and analysis of the low-Ca2+-response plasmid pCD1 of Yersinia pestis KIM5. Infect Immun 66:4611-23
Gehring, A M; DeMoll, E; Fetherston, J D et al. (1998) Iron acquisition in plague: modular logic in enzymatic biogenesis of yersiniabactin by Yersinia pestis. Chem Biol 5:573-86
Hensel, M; Shea, J E; Baumler, A J et al. (1997) Analysis of the boundaries of Salmonella pathogenicity island 2 and the corresponding chromosomal region of Escherichia coli K-12. J Bacteriol 179:1105-11
Blattner, F R; Plunkett 3rd, G; Bloch, C A et al. (1997) The complete genome sequence of Escherichia coli K-12. Science 277:1453-62