genome annotation of the marine bacterium cellulophaga lytica n.
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Genome Annotation of the Marine Bacterium Cellulophaga lytica. Joanna Klein, Ph.D. Northwestern Scholarship Symposium May 4, 2012. What are Bacteria?. Single celled microorganism Friend or Foe? Friend: health, environment, industry Foe: cause a variety of infectious diseases.

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genome annotation of the marine bacterium cellulophaga lytica

Genome Annotation of the Marine Bacterium Cellulophagalytica

Joanna Klein, Ph.D.

Northwestern Scholarship Symposium

May 4, 2012

what are bacteria
What are Bacteria?
  • Single celled microorganism
  • Friend or Foe?
    • Friend: health, environment, industry
    • Foe: cause a variety of infectious diseases
cellulophaga lytica
Cellulophaga lytica
  • Marine bacterium
  • Isolated from beach mud near Limon, Costa Rica in 1969

cellulophaga lytica1
Cellulophaga lytica
  • Gram negative
  • Filamentous
  • Yellow pigmentation
  • Exhibits gliding motility
cellulophaga lytica2
Cellulophaga lytica
  • Member of the Cytophaga-Flavobacterium-Bacteroides (CFB) group of bacteria
    • Poorly characterized branch
phylogenetic tree of bacteria
Phylogenetictree of Bacteria
  • Proteobacteria
    • E. coli, Salmonella, Bordetella, Helicobacter, Vibrio
  • Firmicutes
    • Staphylococcus, Streptococcus, Lactobacillus, Clostridium
cellulophaga lytica3
Cellulophaga lytica
  • Target organism in the Genomic Encyclopedia of Bacteria and Archaea (GEBA) Research Program of the Department of Energy/Joint Genome Institute
  • GEBA organisms
    • 100 representative organisms from each of the branches
    • Organisms with potential energy applications
biofuel production
Biofuel production
  • C. lyticaproduces a variety of enzymes that may have applications in biotechnology and biofuel production
deconstruction by c lytica
Deconstruction by C. lytica
  • C. lyticacontains many polysaccharide degrading enzymes
    • Polysaccharides
      • Large molecules that store energy or provide structure
      • Carbohydrates/starches
      • Cellulose in plant cell walls
    • Enzymes break down polysaccharides into simple sugars that that can be fermented to produce energy
  • Polysaccharide degrading enzymes
    • 3 cellulases
    • 3 fucoidases
    • 1 xylosidase
ethanol production
Ethanol production
  • Ethanol produced as a byproduct of starch degradation and subsequent fermentation
    • Well developed technology
    • Enzymes digest starch into simple sugars which are readily fermented by known microorganisms to produce ethanol
    • Issues…
cellulosic ethanol production
Cellulosic ethanol production
  • Goal is to use the cellulose biomass found in plant cell walls of leaves and wood to produce ethanol
    • Problems to overcome:
      • Lignin, also found in cell wall, hinders digestion of cellulose from wood
      • Enzymes that digest cellulose into simple sugars are poorly understood
      • Organisms that ferment these simple sugars to produce ethanol are poorly understood
  • Can C. lytichelp achieve this goal?
why study c lytica
Why study C. lytica?
  • Model organism to understand the CFB group better
  • Contribute to biofuel research and applications
genome annotation of cellulophaga lytica
Genome Annotation of Cellulophagalytica
  • One way to understand more about the life processes of C. lytica is through a study of its genome.
  • Genome
    • All of the genetic material, DNA, of an organism
    • DNA is made up 4 smaller molecules known as the bases A,C,G &T
sequencing genomes
Sequencing genomes
  • We can easily determine the entire DNA sequence of an organism – it’s genome.
  • DNA sequencing technology has developed rapidly since the human genome project, completed in 2003
    • Took 13 years to complete, involved 100’s of researchers around the globe, and cost a total of of $2.7 billion
    • Entire 3 billion base-pair sequence is available in a public database
genome projects
Genome projects
  • Currently, there are more than 3000 complete or nearly complete genome sequences of microbes available.
  • Over 1200 genome sequencing projects in higher organisms (plants, animals, fungi, protists)
  • The complete genome of Cellulophaga lytica was sequenced by the DOE and published in 2011
    • 3,765,936 bases
computer annotation of c lytica
Computer annotation of C. lytica
  • Number of genes and predicted function of each gene product.



process of annotation
Process of annotation
  • Automatic annotation - done automatically using computer software
      • 35% of computer generated annotations are wrong or are missing information due to limitations of computer algorithms
  • Manual Annotation – humans analyze the information generated by computers and make corrections as necessary.
    • Labor intensive and time consuming
    • Solution: Train students to participate in the process

IMG-ACT is a toolkit of online gene and genome analysis programs.

  • Using IMG-ACT, students annotate genomes
    • provide human expertise necessary for accurate, up-to-date, reliable annotation
  • Students contribute to the scientific community and learn biological concepts through participating in original research
genome annotation of c lytica at nwc
Genome annotation of C. lytica at NWC
  • 39 NWC students have participated in this research endeavor
    • Science Research Institute, Summer 2011
    • Genetics, Fall 2011
    • Microbiology, Spring 2012
  • 15 genes have been fully annotated
  • 10 genes have been partially annotated
restriction endonuclease type i
Restriction endonuclease type I
  • What is the amino acid sequence of the protein encoded by this gene?
  • Used Integrated Microbial Genomes (IMG) database
  • Amy Knight and Allison Lothe
dna topoisomerase iii
DNA topoisomerase III
  • How does this protein compare to the sequence of other proteins?
  • Used BLAST program
  • Libby Nelson and Chelsey Fiecke
rna polymerase sigma subunit 24
RNA polymerase sigma subunit 24
  • What are key functional amino acid residues in the protein?
  • Web Logo Program
  • Silas Baalke and Laura Torgerson
dna replication protein a
DNA Replication Protein A
  • What enzymatic pathway is the protein involved in?
  • Used KEEG Pathway database
  • Marie Abeler and Gabe Jefferson
b galactosidase
  • What pathway is this enzyme found in?
  • KEEG database
  • Daniel Plack,

Michael Lowry

prolyl trna synthase
Prolyl-tRNA synthase
  • What is the 3D structure of similar proteins?
  • ProteinDataBank (PDB)
  • Sarah Ivanca and Victoria Hanson
nusa b g anti termination factors
NusA, B, G anti-termination factors
  • Where is the gene in relation to other genes?
  • Used Gene neighborhood feature of IMG
  • Matt Takata and Zach Fredman
rnase h
RNase H
  • What reaction does the enzyme catalyze?
  • Used Metacyc database
  • Chelsey Fiecke
elongation factor ts
Elongation Factor Ts
  • How closely related is this protein to proteins in other bacteria?
  • Used Phylogeny FR program
  • Ellen Chae, Holly Tomaz
cytochrome c oxidase subunit 3
Cytochrome C oxidase subunit 3
  • Where is this protein located in the cell?
  • TMHMM algorithm
  • Alannah Pratt, Michael Lowry, SRI high school students
  • Are there paralogs of this gene?
  • IMG database query
  • Ryan Bradbury and Luke Delain
rna polymerase sigma 70 factor
RNA polymerase sigma-70 factor
  • Was this gene named properly?
  • Multiple lines of evidence used to change name to RNA Polymerase anti-sigma 70 factor
  • Camaren Terrill and Ben Sorenson
future work
Future work
  • 3,348 genes left to annotate!
    • Special interest in:
      • Polysaccharide degrading enzymes
      • Motility proteins
      • Proteins with unknown function
  • Study the function of interesting genes in the lab
  • NWC students who have participated in this research.
    • Genetics, Microbiology and SRI courses
    • Research students Steven Erickson and Andy Jaeger
  • Northwestern College for providing the opportunity and support for the sabbatical during which this project was initiated.
  • Additional funding received from a 2010-2011 Faculty Development Grant