Jennifer Okonta
From OpenWetWare
Jump to navigationJump to search
Defined Words
- transposases-Transposase is an enzyme that binds to the ends of a transposon and catalyzes the movement of the transposon to another part of the genome by a cut and paste mechanism or a replicative transposition mechanism. en.wikipedia.org/wiki/Transposases
- chemontaxis-movement by a cell or organism in reaction to a chemical stimulus. wordnetweb.princeton.edu/perl/webwn
- prophage-A prophage is a phage genome inserted as part of the linear structure of the DNA chromosome of a bacterium. en.wikipedia.org/wiki/Prophage
- integron-A genetic unit that, among others, encodes proteins that splice gene cassettes into chromosomes, where the cassettes can become functional. www.nature.com/nrg/journal/v5/n3/glossary/nrg1292_glossary.html
- aetiological- the cause of a disease. wordnetweb.princeton.edu/perl/webwn
- heterotrophic- obtaining nourishment from organic substances, not from food produced within the organism.wordnetweb.princeton.edu/perl/webwn
- Transposon- a segment of DNA that can become integrated at many different sites along a chromosome (especially a segment of bacterial DNA that can be translocated as a whole).wordnetweb.princeton.edu/perl/webwn
- pathogenesis- the origination and development of a disease. wordnetweb.princeton.edu/perl/webwn
- Permease - The permeases are membrane transport proteins, a class of multipass transmembrane proteins that facilitate the diffusion of a specific molecule in or out of the cell.en.wikipedia.org/wiki/Permease
- Rho-Independent terminator- The rho-independent signal is found on the DNA template strand and consists of a region that contains a section that is then repeated a few base pairs away in the inverted sequence. http://www.sparknotes.com/biology/molecular/dnatranscription/section2.rhtml
Outline
What is Vibrio Cholerae
- the aetiological agent of cholera
- includes pathognenic and non pathognenic strains that vary in gene content
- contains a wide variety of strains and biotypes
- receives and transfers genes for toxins
- has colonization factors
- is resistant to antibiotics
- has capsular polysaccharides that provide resistance to chlorine and new surface antigens
- it represents a significant portion of the culturable heterotrophic bacteria of oceans, coastal waters and estuaries
- these bacteria strongly influence nutrient cycling in the marine environment
Purpose of the Work
- To determine and analyze the genome sequence of Vibrio Cholerae.
- It is an important step toward the complete molecular description of how this free-living environmental organism became a human pathogen by horizontal gene transfer.
Genome Analysis & Comparative Genome Analysis
- Sequenced by the whole genome random sequencing method.
- Two circular chromosomes make up this genome
- 3,885 predicted open reading frames (ORFs) and 792 predicted Rho-independent terminators
- Most genes required for growth and viability are located on chromosome 1
- two-chromosome structure of V. cholerae allows for comparisons
- There is pronounced asymmetry in the distribution of genes known to be essential for growth and virulence between the two chromosomes
- Chromosome 2 contains a larger fraction (59%) of hypothetical genes and genes of unknown function, compared with chromosome
- Most genes known to be essential in bacterial pathogenicityare located on chromosome 1
Figure Analysis
Figure 1
Linear representation of the V. cholerae chromosomes
- Shows the location of the predicted coding regions, colour-coded by biological role, RNA genes, tRNAs, other RNAs, Rho-independent terminators and Vibrio cholerae repeats.
- Arrows represent the direction of transcription for each predicted coding region
Figure 2
Circular representation of the V. cholerae genome
- 2 chromosomes
- the first and second circles show predicted protein-coding regions on the plus and minus strand
- The third circle shows recently duplicated genes on the same chromosome and on different chromosomes
- The fourth circle shows transposon-related (black), phage-related (blue), VCRs (pink) and pathogenesis genes (red
- The fifth circle shows regions with significant values for trinucleotide composition in a 2,000-bp window
- The sixth circle shows percentage G+C in relation to mean G+C for the chromosome.
- The seventh and eighth circles are tRNAs and rRNAs, respectively.
Figure 3
- Pathways for energy production and the metabolism of organic compounds, acids and aldehydes are shown.
- Transporters are grouped by substrate specificity.
- Question marks associated with transporters indicate a putative gene, uncertainty in substrate specificity, or direction of transport
- Export or import of solutes is designated by the direction of the arrow through the transporter
- Gene location on the two chromosomes, for both transporters and metabolic steps, is indicated by arrow color.
Table 1
General features of the Vibrio cholerae genome.
- self explanatory
- the size, number of ORF, the number of tRNA, the number rRNA, the number of significant and unsignificant proteins and other things are listed.
Figure 4
Percentage of total Vibrio cholerae open reading frames (ORFs) in biological roles compared with other -Proteobacteria
- V. cholerae, chromosome 1 (blue)
- V. cholerae, chromosome 2 (red)
- Escherichia coli (yellow)
- Haemophilus influenzae (pale blue)
Figure 5
Comparison of the V. cholerae ORFs with those of other completely sequenced genomes
- All V. cholerae ORFs (large chromosome, blue; small chromosome, red) were searched against all other genomes with FASTA
Figure 6
Phylogenetic tree of methyl-accepting chemotactic proteins (MCP) homologues in completed genomes
- Amino-acid sequences of the proteins were aligned using CLUSTALW
- Neighbor-joining phylogenetic tree was generated from the alignment using the PAUP program
Methods
- Grown from a single isolated colony
- Cloning, sequencing and assembly were as described for genomes sequenced by TIGR
- Sequences from both ends of served as a genome make up to show the orientation, order and integrity of the contigs
- Sequence gaps were closed by editing the end sequences and/or primer
- The initial set of ORFs was identified with GLIMMER and those shorter than 30 codons were eliminated
- ORFs were searched against a non-redundant protein database
- Frameshifts and point mutations were detected and corrected when needed
- Paralogous gene families were made by searching the ORFs against themselves using the program BLASTX
- Probability values for this analysis are based on the assumption that the DNA composition is relatively uniform throughout the genome
- Homologues of the genes of interest were identified using the BLASTP and FASTA3 search programs
- All homologues were then aligned to each other using the CLUSTALW program with default settings
- Phylogenetic trees were generated from the alignments using the neighbour-joining algorithm
Conclusions
- New starting point for the study of V. Cholerae environmental and pathobiological characteristics
- The genomic sequence of V. cholerae should facilitate the study of this model multi-chromosomal prokaryotic organism
- Might provide important clues to understanding the metabolic and regulatory networks that link genes on the two chromosomes
- Represents a promising genetic system for studying how several horizontally acquired loci located on separate chromosomes can still efficiently interact at the regulatory, cell biology and biochemical levels
Jennifer Okonta 13:32, 17 October 2010 (EDT)