DNA and the hidden world of microbes
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DNA and microbes were the topic of the document. It contained the following key points: 1) Microbes are small but there are lots of them, with more cells on Earth than stars in the universe. They play important roles in ecosystems through processes like nitrogen fixation and the carbon cycle. 2) Studying microbial diversity has progressed through four eras, from identifying the bacterial and archaeal domains using rRNA to metagenomic studies of microbial communities without culturing. 3) Sequencing revolutionized the field by allowing classification of microbes and prediction of their functions from genomes. It revealed novel forms of phototrophy and the vast uncultured majority of microbes.
DNA and the hidden world of microbes
- 1. DNA & Microbes DNA and the Hidden World of Microbes CLIMB Symposium September 12, 2011 Jonathan A. Eisen University of California, Davis
- 2. Robin in London Examples
- 4. MICROBES
- 5. Microbes are small Size * They are small, by definition * Were not really known until microscope invention
- 6. But there are LOTS of them Numbers * 100 million in gram of soil * More cells on Earth than stars in universe * More biomass than plants, animals * 10x cells on humans than human cells * 50x10^6 viruses/ml sea water
- 9. Function 1: The Bad
- 10. Function 2: The Good Nitrogen Fixation Animal Nutrition Carbon Fixation
- 11. Function 3: The Unknown
- 12. Function 4: The Unusual H2S, pH 0, 95°C CO, 80°C High salt low pH CO2 4°C 105°C CH3
- 13. Function 5: Food, Fuel, etc Feed microbes a little carbon and they can make some nice things
- 14. Function 6: Running the Planet Carbon cycle Nitrogen cycle
- 15. Studying microbial diversity • Two main questions • Who is out there? • What are they doing?
- 16. Sequencing and Microbes • Sequencing is useful as a tool in studies of microbial diversity for many reasons • It is complimentary to other means of study • Four major “ERAs” in use of sequencing for microbial diversity studies
- 17. Era I: rRNA Tree of Life
- 18. Era I: rRNA Tree of Life Bacteria • Appearance of microbes not informative (enough) • rRNA Tree of Life Archaea identified two major groups of organisms w/o nuclei • rRNA powerful for many reasons, though not perfect Eukaryotes Barton, Eisen et al. “Evolution”, CSHL Press. 2007. Based on tree from Pace 1997 Science 276:734-740
- 19. Diversity III: Phylogenetic • Three main kinds of organisms • Bacteria • Archaea • Eukaryotes • Viruses not alive, but some call them microbes • Many misclassifications occurred before the use of molecular methods
- 20. The Tree of Life 2006 adapted from Baldauf, et al., in Assembling the Tree of Life, 2004
- 21. The Tree of Life 2006 adapted from Baldauf, et al., in Assembling the Tree of Life, 2004
- 22. Era II: rRNA in environment
- 24. Great Plate Count Anomaly Culturing Microscope Count Count
- 25. Great Plate Count Anomaly Culturing Microscope Count <<<< Count
- 26. Great Plate Count Anomaly DNA Culturing Microscope Count <<<< Count
- 44. CSI Microbiology Collect from environment
- 45. CSI Microbiology Collect from environment
- 46. rRNA PCR DNA extraction PCR Makes lots Sequence PCR of copies of rRNA genes the rRNA genes in sample rRNA1 5’...ACACACATAGGTGGAGC TAGCGATCGATCGA... 3’ Phylogenetic tree Sequence alignment = Data matrix rRNA2 rRNA1 rRNA2 rRNA1 A C A C A C 5’..TACAGTATAGGTGGAGCT rRNA4 AGCGACGATCGA... 3’ rRNA3 rRNA2 T A C A G T rRNA3 rRNA3 C A C T G T 5’...ACGGCAAAATAGGTGGA E. coli Humans rRNA4 C A C A G T TTCTAGCGATATAGA... 3’ Yeast E. coli A G A C A G rRNA4 5’...ACGGCCCGATAGGTGG Humans T A T A G T ATTCTAGCGCCATAGA... 3’ Yeast T A C A G T
- 47. Comparing Sequences • Analogous to comparing bones
- 48. 1992 rRNA PCR and me .... NOTES 3419 A. pisum P A. piswn S Tx. nivea L awaaa sym L equizenata syr L Cud orbgcdar s,ym rs. gesgosterorn - / I -- V I N. gonorrhoeae B. Uhar.opkiuns sym 5% C. magncisca sym Tns. sp. L-12 A. tnefaciens JOURNAL OF BACTERIOLOGY, May 1992, p. 3416-3421 0021-9193/92/103416-06$02.00/0 Vol. 174, No. 10 Copyright © 1992, American Society for Microbiology Phylogenetic Relationships of Chemoautotrophic Bacterial R. ricketsil Symbionts of Solemya velum Say (Mollusca: Bivalvia) Determined by 16S rRNA Gene Sequence Analysis JONATHAN A. EISEN,lt STEVEN W. SMITH,2 AND COLLEEN M. CAVANAUGH`* Department of Organismic and Evolutionary Biology, 1 and Harvard Genome Laboratory,2 Biological Laboratories, Harvard University, Cambridge, Massachusetts 02138 Received 4 November 1991/Accepted 9 March 1992 The protobranch bivalve Solemya velum Say (Mollusca: Bivalvia) houses chemoautotrophic symbionts intracellularly within its gills. These symbionts were characterized through sequencing of polymerase chain rRNA regions and hybridization of an Escherichia coli gene probe to S. velum
- 49. Major phyla of bacteria & archaea No cultures Some cultures
- 50. The Hidden Majority Richness estimates Hugenholtz 2002 Bohannan and Hughes 2003
- 52. Era III: Genome Sequencing
- 53. 1st Genome Sequence Fleischmann et al. 1995
- 54. Genomes Revolutionized Microbiology • Predictions of metabolic processes • Better vaccine and drug design • New insights into mechanisms of evolution • Genomes serve as template for functional studies • New enzymes and materials for engineering and synthetic biology
- 56. Lateral Gene Transfer Perna et al. 2003
- 57. Network of Life Bacteria Archaea Eukaryotes Figure from Barton, Eisen et al. “Evolution”, CSHL Press. Based on tree from Pace NR, 2003.
- 58. Using the Core
- 59. Wh Whole genome tree built using AMPHORA by Martin Wu and Dongying Wu
- 60. Microbial genomes From http://genomesonline.org
- 61. Phylogenetic Diversity • Phylogenetic diversity poorly sampled • GEBA project at DOE- JGI correcting this
- 62. Era IV: Genomes in Environment
- 63. Novel Form of Phototrophy Beja et al. 2000
- 64. Era IV: Genomes in Environment shotgun sequence Metagenomics
- 67. Weighted % of Clones 0 0.1250 0.2500 0.3750 0.5000 Al ph a Be pro ta teo G p b am rot ac m eo te ba ria Ep ap ct si ro lo t e np eob ria D el rot ac ta e t pr ob eria ot ac C eo te ya b r EFG no ac ia EFTu rRNA RecA RpoB b te HSP70 Fi act ria rm e Ac ic ria tin ut es ob a C cte hl r or ia ob C i FB C hl o Major Phylogenetic Group Sp rof Sargasso Phylotypes iro lex i Fu cha D 304: 66. 2004 Metagenomic Phylotyping ei so et no ba es co ct cc er Euus ia ry -T a h Venter et al., Science C rcherm re na aeous rc t ha a eo ta
- 69. Field Diversity
- 71. Generation I: Manual Sanger
- 73. Generation III: No clones
- 75. Acknowledgements • $$$ • DOE • NSF • GBMF • Sloan • DARPA • People, places • DOE JGI: Eddy Rubin, Phil Hugenholtz et al. • UC Davis: Aaron Darling, Dongying Wu • Other: Jessica Green, Katie Pollard, Martin Wu, Tom Slezak, Jack Gilbert
Editor's Notes
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- #6: Small organisms - too small to see without help\n giant microbes\n \n Mostly live as single cells\n \n Many different kinds (more on this in a bit)\n \n VIruses not included by some, but I think they count\n \n (Show flu bug slide)\n \n
- #7: Small organisms - too small to see without help\n giant microbes\n \n Mostly live as single cells\n \n Many different kinds (more on this in a bit)\n \n VIruses not included by some, but I think they count\n \n (Show flu bug slide)\n \n
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- #10: Do lots of nasty things\n Get all the good names by the way b/c mostly known through their diseses\n Yersinia pestis\n Vibrio cholerae\n Bacillus anthracis\n Smallpox - ok not all\n Mycobacterium tuberculosis\n Mycobacterium leprae\n Clostridium tetanus\n Clostridium botulinum\n
- #11: Mutualists (though names not so good)\n N2 fixation\n C fixation - Chloroplasts inside plants are actually symbiotic bacteria\n Digestion - ruminants and all cellulose\n\n
- #12: Cloud of microbes living in / on organisms\n More microbial cells than human\n
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- #15: Microbes run the planet\n All photosynthesis\n Number of microbial cells\n
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- #21: Composite tree of life based on diverse data\nmain point - euks monophyletic\n\nSignif LGT in euks, but mostly bacterial genes\nso bulk of euk genome is vertically transmitted\nand we can use these gene to reconstruct their history\n
- #22: Composite tree of life based on diverse data\nmain point - euks monophyletic\n\nSignif LGT in euks, but mostly bacterial genes\nso bulk of euk genome is vertically transmitted\nand we can use these gene to reconstruct their history\n
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- #42: Many prokaryotes cannot be grown and cannot get them from harsh envts, or replicate the envt (i.e volcanic)\nHow do we approach this problem?\n\nResearcher go to various areas to collect samples from envt\n\nGet samples, put into tube\n
- #43: Send it out for sequencing, do an alignment with your gene and blast it (search for other organisms) with a similar sequence\n
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- #45: Functional prediction using a gene tree is just like predicting the biology of a species using a species tree\n
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- #60: This is a tree of a rRNA gene that was found on a large DNA fragment isolated from the Monterey Bay. This rRNA gene groups in a tree with genes from members of the gamma Proteobacteria a group that includes E. coli as well as many environmental bacteria. This rRNA phylotype has been found to be a dominant species in many ocean ecosystems.\n
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