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MICROBIOLOGIA GENERALE Prokaryotic genomes The Escherichia coli nucleoid The chromosome of Escherichia coli The chromosome of Escherichia coli Autoradiograph of intact replicating chromosome of Escherichia coli The prokaryotic genome Prokaryotes sometimes possess smaller extrachromosomal pieces of DNA called plasmids.The total DNA content of a prokaryote is referred to as the cell genome The prokaryotic genome Core genome versus Pan genome Essential and non-essential genes in the prokaryotic genome Supercoiled DNA Prokaryotic genomes: Sequencing, sizes, ORF contents and gene distributions Whole-genome random sequencing and assembly of Haemophilus influenzae Rd Fleischmann et al. Science 269, 469-512, 1995 An approach for genome analysis based on sequencing and assembly of unselected pieces of DNA from the whole chromosome has been applied to obtain the complete nucleotide sequence (1,830,137 base pairs) of the genome from the bacterium Haemophilus influenzae Rd. This approach eliminates the need for initial mapping efforts and is therefore applicable to the vast array of microbial species for which genome maps are unavailable. The H. influenzae Rd genome sequence (Genome Sequence DataBase accession number L42023) represents the only complete genome sequence from a free-living organism. H. influenzae DNA was sonicated and fragments with sizes between 1.6 and 2.0 kb purified from an agarose gel and ligated into a plasmid vector to produce a clone library. End sequences were obtained from clones taken from this library, and a computer used to identify overlaps between sequences. This resulted in 140 sequence contigs, which were assembled into the complete genome sequence, The steps in a whole-genome shotgun sequencing project How does the computer find an ORF? Locating possible functional open reading frames (ORFs) Escherichia coli genomes: core genome and pathogenic islands Green: genes common to all strains Red: genes present in the pathogenic strains only Blue: genes found only in uropathogen strain 536 How Many Genomes Have Been Sequenced? March 10, 2015 (NCBI) Archaea: 469 Bacteria: 31731 Protists: 266 Animals: 557 Plants: 218 Prokaryotic genomes Microorganism Mycoplasma genitalium Borrelia burgdorferi Size (bp) ORFs 580 070 910 725 470 853 Treponema pallidum 1 138 006 1041 Helicobacter pylori Staphylococcus aureus 1 667 867 2 814 816 1590 2593 Bacillus subtilis Mycobacterium tuberculosis 4 214 810 4 411 529 4100 3924 Escherichia coli Pseudomonas aeruginosa 4 639 221 6 264 403 4288 5570 Streptomyces coelicolor 8 667 507 7846 Total ORFs in genome Correlation between genome size and ORF content in prokaryotes Genome size (megabases) Each megabase of prokaryotic DNA encodes about 1000 ORFs Gene function in bacteria genomes Functional category of genes as a relative percent of the genome versus total ORFs in the genome Variations in gene category in Bacteria and Archea Eukaryotic genomes Organism Saccharomyces cerevisiae Plasmodium falciparum Encephalitozoon cuniculi Size (Mbp) 13 (16 chr) 27 (14 chr) ORFs 5570 - 2.9 (11 chr) 1997 97 (6 chr) 19099 Arabidopsis thaliana Mus musculus 125 (5 chr) 2500 (23 chr) 25498 30000 Homo sapiens 3000 (23 chr) 30000? Caenorhabditis elegans Eukaryotic genomes Bacterial metagenomics How complete microbial genome sequence data can speed vaccine development