Prospects for the Commercial Use of Genetic Engineering in Biofuel Production

David J. Glass, Ph.D.D. Glass Associates, Inc.

Presented at EUEC 2010, Phoenix, AZFebruary 2, 2010

Overview: Prospects for Use of Advanced Biotechnology for Biofuel ProductionSummary of technology strategies.Companies applying advanced biotechnology to

biofuels.Impact of biotechnology regulations on biofuels.Commercialization status and prospects.

Technology Strategies

Biotechnologies Applicable to BiofuelsClassical microbial mutation and selectionPlant breedingRecombinant DNADirected evolutionDNA shufflingMetabolic engineeringSynthetic biology

Genetic Engineering Strategies: Microorganisms• Overexpress desired enzymes (e.g. to improve

ability to process or degrade cellulosic feedstocks).

• Engineer microorganisms to manufacture novel or improved industrial enzymes.

• Create novel or synthetic microorganisms or new metabolic pathways to enable production of renewable fuels.

Genetic Engineering Strategies: AlgaeEnhance algal growth rate.Enhance or alter lipid biosynthesis.Enhance photosynthesis.Enable use of alternate

food sources.Enable secretion of lipids

to aid oil/water separation.

Genetic Engineering Strategies: PlantsOverexpress cell-wall hydrolysis enzymes.

Cellulases, Hemicellulases, Ligninases.Increase plant biomass, cellulosic biomass.

Cellulose biosynthetic enzymes.Lignin modification to reduce need for

pretreatment.Down-regulate lignin biosynthesis.

Regulated gene expression, so traits are active only when needed.

Companies applying advanced biotechnology to biofuels

Companies Using or Developing Modified Microorganisms: EthanolBioEnergy InternationalDuPont DaniscoGlycos BiotechnologiesGreenTech AmericaJoule BiotechnologiesMascoma

Microbiogen (non-GMO)

PromethegenQterosTMO RenewablesVereniumZymetis

Note: Several of these companies are also developing other fuels.

Companies Using or Developing Modified Microorganisms: Biodiesel, Other FuelsBiodiesel, petroleum fuelsAmyris BiotechnologiesCodexisGlycos BiotechnologiesJoule BiotechnologiesLS9PromethegenSequescoSynthetic Genomics

Butanol, IsobutanolBioenergy InternationalButamax (DuPont/BP)Cobalt BiofuelsGevoGreen Biologics, Ltd.METabolic ExplorerTetraVitae Bioscience

Companies Using Newer Microbial TechnologiesAmyris Biotechnologies: synthetic biologyCodexis: DNA shufflingGlycos Biotechnologies: metabolic engineeringJoule Biotechnologies: synthetic biologyLS9: synthetic biologySynthetic Genomics: synthetic biologyTMO Renewables: thermophilic

microorganismsVerdezyne: synthetic biology,

metabolic engineering

Companies Manufacturing Enzymes for Biofuels using Modified OrganismsMicroorganismsAB EnzymesDanisco/GenencorRoyal DSM N.V.Dyadic InternationalIogen CorporationNovozymes A/SProtéus

PlantsInfinite Enzymes

(transgenic plants)Medicago

(transient expression)Syngenta

(transgenic plants)

Companies Reported to be Developing Genetically Modified Algae for BiofuelsAlgenol BiofuelsAurora BiofuelsAXI LLCGlobal Green SolutionsSapphire EnergySolazymeSynthetic GenomicsTargeted Growth

Note: companies included based on press, Internet reports: use of GMOs not necessarily confirmed on company websites. Not all these companies are necessarily using genetic engineering.

Companies Developing Transgenic Plants for BiofuelsConventional FeedstocksAgrivida: cornArborGen: purpose-grown treesEdenspace Systems: cornTargeted Growth: corn

Note: some of these companiesare also developing newer feedstocks.

Companies Developing Transgenic Plants for Biofuels

Newer FeedstocksAgragen: CamelinaAgrisoma: Brassica and JatrophaAgrivida: switchgrass, sugarcane,

sorghum, othersCeres: non-food grassesEdenspace Systems: switchgrassEvogene: canola, soybean, others

Companies Developing Transgenic Plants for Biofuels

Newer FeedstocksFarmacule BioIndustries: sugarcane, tobaccoMendel Biotechnology: grasses, othersMetabolix: switchgrass, oil crops, othersSG Biofuels: JatrophaTargeted Growth: Camelina,

canola, others

Impact of Biotechnology Regulations on Biofuels

Overview of U.S. Regulation of Environmental BiotechnologyEnvironmental Protection AgencyMicrobial pesticides.Plant pesticides (e.g. transgenic plants).Engineered microorganisms not regulated by

other agencies (e.g. nitrogen fixation, bioremediation, industrial applications).

U.S. Department of AgricultureTransgenic plants, including plants producing

industrial or pharmaceutical products.Transgenic animals.

EPA Regulation of Engineered MicroorganismsEPA regulations under the Toxic Substances Control Act

(TSCA) cover industrial uses of microorganisms not regulated by other agencies.

Applies to “intergeneric” organisms for industrial enzyme production, biotreatment, other bioprocessing uses.

Applicability to algae not certain.Submit Microbial Commercial Activity Notice (MCAN) with

data package 90 days before starting commercial use.Most research and pilot projects are not regulated.Some exemptions available for commercial use.

USDA Regulation of Transgenic PlantsRegulations issued in 1987 cover outdoor uses of

most transgenic plants.Rules have been continually relaxed, most

agricultural uses require only agency notification.Starting in 2003, permits now required for industrial,

pharmaceutical, phytoremediation applications.Permit applications: must be submitted 120 days in

advance of proposed field use.Commercialization via “delisting” petition process.USDA currently considering revisions to regulations.

Regulation of Engineered AlgaeJurisdiction of EPA, USDA under biotech regulations

unclear.Proposed use of engineered algae in pharmaceutical

production in Hawaii in 2005 generated controversy, no federal agency claimed jurisdiction: state law and state courts ultimately determined outcome.

2008 USDA opinion letter to Coastal BioMarine stated no USDA oversight over engineered algal strain if no “plant pest” sequences, and no oversight for smaller-scale use in contained reactors.

Reactor design may be critical in determining regulatory status.

Impact of Biotechnology RegulationsWould not necessarily affect all biofuel organisms,

exemptions may be available.For those projects affected, some additional

activities and expense may be required.Straightforward path to commercial approval,

with many prior successes in agricultural, industrial biotechnology.

The regulatory process can be successfully managed.

Commercialization Status and Prospects

Commercial Status of Modified Microorganisms in BiofuelsVerenium: only MCAN reviewed by EPA under

TSCA for modified ethanol production strain.Demonstration or pilot plants in operation or under

construction: Ethanol production: BioEnergy, DuPont Danisco,

Iogen, Mascoma, Qteros, Verenium.Gevo: demonstration butanol plant.Amyris: pilot plant for petroleum-equivalent fuels.Pilot or commercial algae plants: Algenol, Aurora,

Sapphire, Solazyme, Solix (not necessarily using engineered algae).

Field Uses of Transgenic Plants for BiofuelsSyngenta: Approval for commercial use of amylase-

expressing corn in several countries, approval pending in U.S. since 2005.

Agrisoma: field trials in Canada of engineered Brassica and soybean, improved in oil quality, content and seed size for biodiesel use, 2009.

Infinite Enzymes, Edenspace, Targeted Growth, ArborGen: field trials in the U.S. of various species, including tobacco, corn, Eucalyptus, expressing biodegradative enzymes, 2006-09.

Notable Business Deals Involving Advanced Biotechnology for BiofuelsBP/Martek: Conversion of sugars to biodiesel.BP/Verenium: Cellulosic biofuel development.Chevron/Solazyme: Biodiesel production from algae.Dow Chemical/Algenol: Pilot plant for algal production of

ethanol.ExxonMobil/Synthetic Genomics: Synthetic algal strains

for biofuel production.Royal Dutch Shell/Codexis: Novel methods for

conversion of non-food biomass to biofuels.Royal Dutch Shell/HR Petroleum: marine algae for

biofuels.

Prospects for the Future: Commercialization ChallengesUse of modified organisms will face the same challenges as other biofuel applications:Scale-up to commercial scale.Process economics.Competition from other biological or physical

methods of biofuel manufacture.Feedstock availability and cost; conflict with food

use of cropsAvailability of sufficient land to grow feedstocks or to

build reactors.

Prospects for the Future: Commercialization ChallengesBut use of modified organisms will also face unique challenges:Technical hurdles to accomplish performance goals.Recovering the high R&D costs of advanced

biotech.Government policies, including biotechnology

regulation, especially outside the U.S.Public acceptance of biotechnology solutions.

Prospects for the Future: Commercialization StrategiesIntegration of biological processes with other

downstream process improvements.Diversification:

Augment energy production by co-production of value-added end-products: specialty chemicals, bioplastics.

Entry of tree, plant genetics companies into biofuels.Carbon capture: design systems to capture CO2

streams for use as biofuel feedstock.Novel feedstocks: use of cellulosic waste, novel

nonfood crop species as biofuel feedstocks.

Thank you very muchDavid J. Glass, Ph.D.

D. Glass Associates, Inc.

124 Bird Street

Needham, MA 02492

Phone 617-653-9945

DGlassAssc@aol.com

www.dglassassociates.com

More detailed information on the subjects presented in this talk can be found at my blog:

http://dglassassociates.wordpress.com