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Brazil’s Bioethanol (& GtL Economic modeling needs)

DOE GTL Vertically Integrated BioEnergy Research Center (special thanks to Harvard Inst. for Biologically Inspired Engineering). Brazil’s Bioethanol (& GtL Economic modeling needs). Land use:45,000 km² Sugarcane: 344 million tons (76 tons/ha) Sugar: 23 million tons

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Brazil’s Bioethanol (& GtL Economic modeling needs)

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  1. DOE GTL Vertically Integrated BioEnergy Research Center(special thanks to Harvard Inst. for Biologically Inspired Engineering)

  2. Brazil’s Bioethanol(& GtL Economic modeling needs) Land use:45,000 km² Sugarcane: 344 million tons (76 tons/ha) Sugar: 23 million tons Ethanol:14 million m³ $0.26/L (feedstock 70%) yield increase 3.5%/yr Dry bagasse: 50 million tons Electricity: 1350 MW Bagasse ash 2.5% (vs 40% for coal), nearly no sulfur. Burns at low temperatures, so low nitrogen oxides. Saccharum officinarum

  3. The ‘G’ in GtL '77 Gilbert & Sanger papers '77 developed 1st auto-sequence-reading software '77 1st full plasmid sequence (recombinant DNA) '84 ‘Genomic Sequencing’ PNAS paper & DOE Alta meeting '87 1st Genome grant (DOE) '90 co-PI on 1st NIH Genome Centers, Stanford,GTC,MIT (now Broad) '94 1st genome sequence H.pylori 1.7 Mbp (commercial) '02 Only DOE Center to address all 5 GtL goals '06 SynBERC grant with LBL, MIT, etc. #1: Protein complexes & Mass Spec #2: Regulatory Networks & RNA #3: Microbial Communities #4: Computational models #5: Synthetic Biology

  4. Vertical Integration (horizontal options) PlantsZea,Miscanthus,Poplar, Crambe,Algae Pretreat acid, NH4 fiber explosion (AFEX), silage SaccharifyTrichoderma, Clostridia, Aspergillus Soluble, cellulosome, consolidated FermentSaccharomyces, Escherichia, Zymomonas, Pichia, Klebsiella, communities Extract distill, gas strip, zeolite, phase Process crack, blend ethanol, butanol, ethylesters, alkanes, H2 Uses  transportation, chemicals, power AGRICULTURE Enabling Tech: Modeling, Synthesis, Evolution, Sequencing

  5. Our GTL network Ingram-UFL Celunol Cerrina-UW Endy Codon Devices Collins-BU Chisholm-MIT LS9 Polz DuPont Shell Church-Harvard Silver Shih SynBERC-iGEM BioEnergy International Richard-PSU Agrivida Ausubel-MGH Mendel Hamilton-INL Liu-BNL Curtis-PSU Chromatin Chappell-UKY GreenFuel Laible-ANL Pendse-UME SunEthanol Leschine-UMA Vance-WashU

  6. Enabling Technologies #1: Computational & Systems Modeling

  7. Enabling Technologies #2:Lab evolution (building on systems design)#3: Functional metagenomics (interspecies DNA transfer) Radiation resistance (Edwards & Battista) Tyr/Trp production & transport (Lin & Reppas) Citrate utilization (Lenski) Lactate production (Ingram) Temperature/acid tolerance (Marliere) Glycerol utilization (Palsson) Aldehyde resistance (Sommer & Dantas)

  8. Whole-Genome resequencing of evolved DTrp ompF – non specific transport channel • Glu-117 → Ala (in the pore) • Charged residue known to affect pore size and selectivity • Can increase import & export capability simultaneously Shendure, et al. (2005) Science309:1728

  9. Tech #4: ‘Next Generation’ Sequencing Multi-molecule Reaction Volume AB/APG Ligase beads 1 fL 454/Roche Pol beads 100,000 fL Solexa Pol term 1 fL CGI Ligase 1 fL Affymetrix Hybr array 100 fL IBS Pol beads 10,000 fL Single molecules Helicos Biosci Pol <1fL Visigen Biotech Pol FRET <1fL Pacific Biosci Pol <1fL Agilent Nanopores <1fL fL =1E-15 liters (femto) Our lab has been involved in 8/10

  10. #5: Sequencing genomes from single cells (single chromosome, cell , RNA or particle) Zhang, et al. (2006) . Nature Biotech. June ’06 1) Environmental samples (poor or no lab growth) 2) Candidate chromosome region sequencing 3) Prioritizing or pooling (rare) species based on an initial DNA screen (metagenomics) 4) Multiple chromosomes in a cell or virus 5) RNA 5’, 3’ ends & splicing 6) Cell-cell interactions (predator-prey, symbionts, commensals, parasites) Phi-29 Polymerase Stand-displacement amplification

  11. #7: Recombinational Genome Engineering #6: 10 Mbp of DNA / $1000 chip Digital Micromirror Array 8K Atactic/Xeotron/Invitrogen Photo-Generated Acid 12K Combimatrix/Codon Electrolytic 44K Agilent Ink-jet standard reagents 380K Nimblegen/GA Photolabile 5'protection Amplify pools of 50mers using flanking universal PCR primers & 3 paths to 10X error correction Tian et al. Nature. 432:1050; Carr & Jacobson 2004 NAR; Smith & Modrich 1997 PNAS

  12. #8: 3D Structural Biology – especially membrane and fibrous structures Shih, Douglas, iGEM Hanson & Laible #9, 10: Proteomics, Metabolomics

  13. DOE GTL Vertically Integrated Bioenergy and Novel Technologies(VIBRANT)

  14. How do we distinguish ourselves from other GTL proposals? The DOE-GTL-BRC competition (2 – 3 will be funded) 1. LBL-LLNL-Sandia-Stanford-U.IL 2. ORNL-NREL-Dartmouth-Noble-OK 3. Harvard-MIT-UMA-UME-BU-UFL-PSU-UWI-ANL-INL-BNL- 11 Companies 4. San Diego-Iowa-JCVI 5. Purdue-MIT2 6. UWI-MSU 7. LANL BP competition (1 will be funded) 1. Imperial College, UK 2. Cambridge, UK 3. LBL-LLNL-Sandia-Stanford 4. San Diego-Iowa-JCVI 5. MIT-Purdue

  15. Snapshots from the 1st DOE-VIBRANT meeting Boston 21-Nov-2006

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