Biofuels from algae: Challenges and potential

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

The challenges to algal biofuels include strain identification and improvement in oil productivity and crop protection, nutrient and resource allocation and use, and production of co-products to improve the economics of the entire system.

“These challenges include strain identification and improvement, both in terms of oil productivity and crop protection, nutrient and resource allocation and use, and the production of co-products to improve the economics of the entire system.”

Classified as probably not a substantial claim. May combine two claims.

Algae biofuels may provide a viable alternative to fossil fuels, but the technology must overcome a number of hurdles before it can compete in the fuel market and be broadly deployed.

“Algae biofuels may provide a viable alternative to fossil fuels; however, this technology must overcome a number of hurdles before it can compete in the fuel market and be broadly deployed.”

May combine two claims.

Estimated costs of a barrel of algae-based fuel using current technology are US$300-2600, compared with $40-80 (2009) for petroleum.

“estimated costs of a barrel of algae-based fuel using current technology is US$300–2600, compared with $40–80 (2009) for petroleum [9–12].”

• cites [9] Dismukes GC Carrieri D Bennette N Ananyev GM Posewitz MC Aquatic phototrophs: efficient alternatives to land-based crops for biofuels Curr Opin Biotechnol 19 3 235 240 2008 18539450 sourceNot checked yet

  Not checked in the free run.

• cites [10] Borowitzka MA Algal biotechnology products and processes – matching science and economics J Applied Phycology 4 267 279 1992 sourceNot checked yet

  Not checked in the free run.

• cites [11] Chisti Y Biodiesel from microalgae Biotechnol Adv 25 3 294 306 2007 17350212 sourceNot checked yet

  Not checked in the free run.

• cites [12] Alabi AO Tampier M Bibeau E Microalgae Technogies and Processes for Biofuels/Bioenergy Production in British Columbia The BC Innovation Council BC, Canada 2009Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Iron is required for electron transport in all known photosynthetic organisms and is typically found in iron-sulfur clusters in a variety of photosynthetic proteins.

“Biologically, iron is required for electron transport in all known photosynthetic organisms, including Chlamydomonas reinhardtii, and is typically found in iron-sulfur clusters in a variety of photosynthetic proteins [50].”

• cites [50] Godman J Balk J Genome analysis of chlamydomonas reinhardtii reveals the existence of multiple, compartmentalized iron-sulfur protein assembly machineries of different evolutionary origins Genetics 179 1 59 68 2008 18493040 PMC2390635 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Providing a cheap source of fixed nitrogen will be important for algae biofuel production, and nitrogen-fixing cyanobacteria may help minimize these costs.

“Providing a cheap source of fixed nitrogen will be important for algae biofuel production, and the possibility of using nitrogen-fixing cyanobacteria to supply this nitrogen may help minimize these costs [46].”

• cites [46] Berman-Frank I Lundgren P Falkowski P Nitrogen fixation and photosynthetic oxygen evolution in cyanobacteria Res Microbiol 154 3 157 164 2003 12706503 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Algae require sources of phosphorus, nitrogen and potassium, which are major components of agricultural fertilizers, and large-scale aquaculture will impact these already limited supplies.

“Algae, similar to plants, require sources of phosphorus, nitrogen and potassium, which are the major components of agricultural fertilizers, and large-scale aquaculture will impact these already limited supplies.”

May combine two claims.

Crude algae oil is chemically similar to crude fossil fuel oil, so the engineering challenges associated with algae oil conversion to usable liquid fuels are similar to those already managed by petroleum companies, although improved catalysts will be required to improve gasoline production from bio-oil.

“Once extracted, because crude algae oil is chemically similar to crude fossil fuel oil, the engineering challenges associated with algae oil conversion to usable liquid fuels are similar to those already well managed by petroleum companies, although improved catalysts will be required to improve gasoline production from bio-oil [31].”

• cites [31] Maher KD Bressler DC Pyrolysis of triglyceride materials for the production of renewable fuels and chemicals Bioresour Technol 98 12 2351 2368 2007 17166713 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Algae grown in open ponds have water requirements per unit area similar to cotton or wheat, but less than corn, to replenish water lost in evaporation.

“Perhaps surprisingly, algae grown in open ponds have water requirements per unit area similar to that of cotton or wheat, but less than that of corn, to replenish the water lost in evaporation (for an overview of water requirements of terrestrial plants used in biofuel production see [33]).”

• cites [33] Gerbens-Leenes W Hoekstra AY Van Der Meer TH The water footprint of bioenergy Proc Natl Acad Sci USA 106 25 10219 10223 2009 19497862 PMC2690604 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Sulfur is required for protein synthesis and lipid metabolism, in addition to its key role in the electron transport chain.

“Sulfur, in addition to its key role in the electron transport chain, is also required for protein synthesis and lipid metabolism.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

Iron in its oxidized form is not optimal for uptake, and most algae prefer chelated iron.

“Iron in its oxidized form is not optimal for uptake, and most algae prefer chelated iron.”

May combine two claims.

Photobioreactors have advantages over open systems in that they can more easily maintain axenic cultures and more controlled growth environments, which may lead to increases in productivity; however, contained systems are challenged by efficiencies in gas exchange and a requirement for supplemental cooling.

“PBRs have advantages over open systems in that they can more easily maintain axenic cultures, and can maintain more controlled growth environments, which may lead to increases in productivity; however, contained systems are challenged by efficiencies in gas exchange and a requirement for supplemental cooling [28].”

• cites [28] Chisti Y Biodiesel from microalgae beats bioethanol Trends Biotechnol 26 3 126 131 2008 18221809 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Algae can produce biomass very rapidly, with some species doubling in as few as 6 h, and many exhibiting two doublings per day.

“Algae can produce biomass very rapidly, with some species doubling in as few as 6 h, and many exhibiting two doublings per day [16,17].”

• cites [16] Sheehan J Dunahay T Benemann J Roessler P A Look Back at the U S Department of Energy's Aquatic Species Program – Biodiesel from Algae National Renewable Energy Laboratory CO, USA 328 1998 ▪ ▪ Outlines some of the early successes and challenges faced by groups currently addressing algae biofuels. sourceNot checked yet

  Not checked in the free run.

• cites [17] Huesemann MH Hausmann TS Bartha R Aksoy M Weissman JC Benemann JR Biomass productivities in wild type and pigment mutant of Cyclotella sp. (diatom) Appl Biochem Biotechnol 157 3 507 526 2009 18597048 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Micro- and macro-nutrient supplements, or fertilizer, account for significant costs in the current terrestrial agriculture industry, and biofuels are not expected to be an exception.

“Micro- and macro-nutrient supplements, or fertilizer, account for significant costs in the current terrestrial agriculture industry [38], and biofuels are not expected to be an exception.”

• cites [38] US Department of Agriculture Farm Production Expenditures 2008 Summary US Department of Agriculture DC, USA 80 2009Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

Some estimates for a barrel of algae oil in specific regions reach as low as $84.

“Although some estimates for a barrel of algae oil in specific regions reach as low as $84 [13].”

• cites [13] Huntley ME Redalje DG CO2 mitigation and renewable oil from photosynthetic microbes: a new appraisal Mitigat Adapt Strategies Global Change 12 573 608 2006 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Production of algae biofuels would minimize land use compared with biofuels produced from terrestrial plants, and waste streams can be remediated during culturing of the microalgae.

“Thus, not only would production of algae biofuels minimize land use compared with biofuels produced from terrestrial plants but, in the process of culturing these microalgae, waste streams can be remediated.”

• AssumptionRests on an assumption: Algae can be grown on land that would not be used for traditional agriculture.
• AssumptionRests on an assumption: Algae are very efficient at removing nutrients from water.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Optimal growth of many algal species requires chelated iron and sulfur.

“In addition, optimal growth of many algal species requires chelated iron and sulfur.”

May combine two claims.

Algae efficiently use CO2 and are responsible for more than 40% of global carbon fixation, with the majority of this productivity coming from marine microalgae.

“Algae efficiently use CO2, and are responsible for more than 40% of the global carbon fixation, with the majority of this productivity coming from marine microalgae [14,15].”

• cites [14] Falkowski PG Barber RT Smetacek VV Biogeochemical controls and feedbacks on ocean primary production Science 281 5374 200 207 1998 9660741 sourceNot checked yet

  Not checked in the free run.

• cites [15] Parker MS Mock T Armbrust EV Genomic insights into marine microalgae Annu Rev Genet 42 619 645 2008 18983264 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based).

Many nonarable regions have substantial alkaline or saline water reservoirs beneath them, providing a significant source of nonpotable water suitable for growth of many algal species.

“Expansion of algal growth into nonarable land will require water; fortunately, many of these regions have substantial alkaline or saline water reservoirs beneath them, providing a significant source of nonpotable water that is suitable for growth of many algal species.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

Terrestrial models for large-scale algal aquaculture use land that is not presently used for food agriculture and has minimal known environmental or other significant economic utility.

“The terrestrial models use land that is not presently used for food agriculture, and has minimal known environmental or other significant economic utility.”

May combine two claims.

These oil extraction technologies have all been successfully demonstrated but are relatively expensive in terms of equipment needed or energy required to extract the oil.

“These technologies have all been successfully demonstrated but are relatively expensive, either in terms of equipment needed or energy required to extract the oil.”

• cites [30] Krichnavaruk S Shotipruk A Goto M Pavasant P Supercritical carbon dioxide extraction of astaxanthin from Haematococcus pluvialis with vegetable oils as co-solvent Bioresour Technol 99 13 5556 5560 2008 18068354 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Some cyanobacteria can fix nitrogen, while almost all algal species identified to date require an exogenous source of fixed nitrogen, and most prefer ammonia.

“Some cyanobacteria also have the ability to fix nitrogen, while almost all algal species identified to date require an exogenous source of fixed nitrogen, and most prefer ammonia, as it is less energetically demanding than nitrate or nitrite [45].”

• cites [45] Inokuchi R Kuma Ki Miyata T Okada M Nitrogen-assimilating enzymes in land plants and algae: phylogenic and physiological perspectives Physiol Plant 116 1 1 11 2002 12207656 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Potential waste streams for algal culture include municipal wastewater to remove nitrates and phosphates before discharge, and flue gas of coal or other combustible-based power plants to capture sulfates and CO2.

“Potential waste streams include municipal wastewater to remove nitrates and phosphates before discharge, and flue gas of coal or other combustible-based power plants to capture sulfates and CO2 [22–24].”

• cites [22] He P Xu S Zhang H Bioremediation efficiency in the removal of dissolved inorganic nutrients by the red seaweed, Porphyra yezoensis, cultivated in the open sea Water Res 42 4–5 1281 1289 2008 17959220 sourceNot checked yet

  Not checked in the free run.

• cites [23] Fierro S Sanchez-Saavedra Mdel P Copalcua C Nitrate and phosphate removal by chitosan immobilized Scenedesmus Bioresour Technol 99 5 1274 1279 2008 17531478 sourceNot checked yet

  Not checked in the free run.

• cites [24] Douskova I Doucha J Livansky K Simultaneous flue gas bioremediation and reduction of microalgal biomass production costs Appl Microbiol Biotechnol 82 1 179 185 2009 19096837 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

Addition of iron to induce an algal bloom has been considered and tested as a strategy to sequester CO2.

“Interestingly, the addition of iron to induce an algal bloom has been considered and tested as a strategy to sequester CO2 [47–49].”

• cites [47] Coale KH Johnson KS Chavez FP Southern ocean iron enrichment experiment: carbon cycling in high- and low-si waters Science 304 5669 408 414 2004 15087542 sourceNot checked yet

  Not checked in the free run.

• cites [48] Boyd PW Law CS Wong CS The decline and fate of an iron-induced subarctic phytoplankton bloom Nature 428 6982 549 553 2004 15058302 sourceNot checked yet

  Not checked in the free run.

• cites [49] Coale KH Johnson KS Fitzwater SE A massive phytoplankton bloom induced by an ecosystem-scale iron fertilization experiment in the equatorial pacific ocean Nature 383 6600 495 501 1996 18680864 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Higher dollar estimates for algae fuel are more common and similar to the authors' own estimates, and exclude algal oil from the current liquid fuel market.

“The higher dollar estimates are more common and similar to our own estimates, and exclude algal oil from the current liquid fuel market.”

May combine two claims.

Algae production strains have the potential to be bioengineered, allowing improvement of specific traits and production of valuable co-products, which may allow algal biofuels to compete economically with petroleum.

“Algae production strains also have the potential to be bioengineered, allowing improvement of specific traits [25,26] and production of valuable co-products, which may allow algal biofuels to compete economically with petroleum.”

• cites [25] Rosenberg JN Oyler GA Wilkinson L Betenbaugh MJ A green light for engineered algae: redirecting metabolism to fuel a biotechnology revolution Curr Opin Biotechnol 19 5 430 436 2008 18725295 sourceNot checked yet

  Not checked in the free run.

• cites [26] Zaslavskaia LA Lippmeier JC Shih C Ehrhardt D Grossman AR Apt KE Trophic conversion of an obligate photoautotrophic organism through metabolic engineering Science 292 5524 2073 2075 2001 11408656 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Nitrogen is not limited in supply but is often a limiting macronutrient for plant and algae growth.

“Nitrogen, unlike phosphorous, is not limited in supply but is often a limiting macronutrient when it comes to plant and algae growth.”

May combine two claims.

Large algal monocultures will be invaded by pests and pathogens, so crop protection is a major challenge to algal pond sustainability.

“Much like terrestrial monocultures, large algal monocultures will be invaded by pests and pathogens, and therefore, crop protection is a major challenge to algal pond sustainability.”

May combine two claims.

The Chlorella ellipsoidea nuclear genome was engineered to produce rabbit neutrophil peptide-1, which proved to be effective against human pathogens in vitro.

“The Chlorella ellipsoidea nuclear genome was engineered to produce rabbit neutrophil peptide-1, which proved to be effective agent against human pathogens in vitro [79].”

• cites [79] Chen Y Wang Y Sun Y Zhang L Li W Highly efficient expression of rabbit neutrophil peptide-1 gene in chlorella ellipsoidea cells Curr Genet 39 5 365 370 2001 11525411 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

The aforementioned proteins might be effective against microorganisms, but aquatic invertebrates could still feed freely on these algae.

“The aforementioned proteins might be effective against microorganisms but aquatic invertebrates could still feed freely on these algae.”

May combine two claims.

Insecticidal proteins similar to those from Bacillus thuringensis and Bacillus sphaericus have been expressed in plants and cyanobacteria and shown to be detrimental towards aquatic insects, aquatic larvae and daphnids.

“These proteins have already been expressed in plants and cyanobacteria, and shown to be detrimental towards aquatic insects, aquatic larvae and daphnids [73,83–85].”

• cites [73] Berry JP Gantar M Perez MH Berry G Noriega FG Cyanobacterial toxins as allelochemicals with potential applications as algaecides, herbicides and insecticides Mar Drugs 6 117 146 2008 18728763 PMC2525484 sourceNot checked yet

  Not checked in the free run.

• cites [83] Kuci Ska J Lonc E Rydzanicz K Transgenic bioinsecticides inimical to parasites, but imical to environment Wiadomo Ci Parazytologiczne 49 1 11 2003 16889013Not checked yet

  Not checked in the free run.

• cites [84] Rosi-Marshall Ej Tank Jl Royer Tv Toxins in transgenic crop byproducts may affect headwater stream ecosystems Proc Natl Acad Sci USA 104 41 16204 2007 17923672 PMC2042185 sourceNot checked yet

  Not checked in the free run.

• cites [85] Bøhn T Primicerio R Hessen DO Traavik T Reduced fitness of Daphnia magna fed a bt-transgenic maize variety Arch Environment Contamin Toxicol 55 4 584 592 2008 18347840 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Transformed Chlamydomonas reindhartii chloroplast accumulated 5% total soluble protein of the mammary-associated serum amyloid A3 peptide, but its antimicrobial activity was not tested.

“Transformed Chlamydomonas reindhartii chloroplast accumulated 5% total soluble protein (TSP) of the mammary-associated serum amyloid A3 peptide, but its antimicrobial activity was not tested [80,81].”

• cites [80] Manuell AL Beligni MV Elder JH Robust expression of a bioactive mammalian protein in chlamydomonas chloroplast Plant Biotechnol J 5 3 402 412 2007 17359495 sourceNot checked yet

  Not checked in the free run.

• cites [81] Molenaar AJ Harris DP Rajan GH The acute-phase protein serum amyloid a3 is expressed in the bovine mammary gland and plays a role in host defence Biomarkers 14 1 26 37 2009 19283521 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Outdoor pond systems have lower initial capital costs, but historically these open pond systems have relied mainly on outcompeting contaminating organisms by using densely grown axenic or nearly axenic starter cultures.

“Outdoor pond systems have lower initial capital costs, but historically these open pond systems have relied mainly on outcompeting contaminating organisms by using densely grown axenic (or nearly axenic) starter cultures [58,59].”

• cites [58] Lebeau T Robert JM Diatom cultivation and biotechnologically relevant products. Part I. Cultivation at various scales Appl Microbiol Biotechnol 60 6 612 623 2003 12664139 sourceNot checked yet

  Not checked in the free run.

• cites [59] Matsudo MC Bezerra RP Sato S Perego P Converti A Carvalho JCM Repeat fed-batch cultivation of Arthrospira (spirulina) patensis using urea as nitrogen source Biochem Engineer J 43 1 52 57 2008 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Expression of bovine lactoferricin was achieved in nuclear transformants of Nannochloropsis oculata, and algal extracts from strains expressing this protein were effective against Escherichia coli and Vibrio parahemolyticus.

“Recently, expression of bovine lactoferricin was achieved in nuclear transformants of Nannochloropsis oculata. Algal extracts from strains expressing this protein were effective against Escherichia coli and Vibrio parahemolyticus [82].”

• cites [82] Li SS Tsai HJ Transgenic microalgae as a non-antibiotic bactericide producer to defend against bacterial pathogen infection in the fish digestive tract Fish Shellfish Immunol 26 2 316 325 2009 18691655 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Dunaliella salina and Arthrospira can withstand up to 35% salinity and pH 10, respectively.

“This would be the case with Dunaliella salina and Arthrospira, which can withstand up to 35% salinity and pH 10, respectively [59–61].”

• cites [59] Matsudo MC Bezerra RP Sato S Perego P Converti A Carvalho JCM Repeat fed-batch cultivation of Arthrospira (spirulina) patensis using urea as nitrogen source Biochem Engineer J 43 1 52 57 2008 sourceNot checked yet

  Not checked in the free run.

• cites [60] Ginzburg M Ginzburg BZ Interrelationships of light, temperature, sodium chloride and carbon source in growth of halotolerant and halophilic strains of Dunaliella Eur J Phycology 16 3 313 324 1981Not checked yet

  Not checked in the free run.

• cites [61] Dubinsky Z Rotem J Relations between algal populations and the pH of their media Oecologia 16 53 69 1974 28308951 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Closed systems such as photobioreactors have the potential to minimize contamination but at a high capital expense.

“Closed systems, such as PBRs, have the potential to minimize contamination, but this comes at a high capital expense.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The majority of antibiotic extracts studied so far have been from marine macro- and micro-algae, but they are also present in many freshwater species.

“The majority of antibiotic extracts studied so far have been from marine macro- and micro-algae [65–67]; however, they are also present in many freshwater species [67–69].”

• cites [65] Bhadury P Wright PC Exploitation of marine algae: biogenic compounds for potential antifouling applications Planta 219 4 561 578 2004 15221382 sourceNot checked yet

  Not checked in the free run.

• cites [66] Pesando D Antibacterial and antifungal activities of marine algae Intro Applied Phycol 3 26 1990Not checked yet

  Not checked in the free run.

• cites [67] Gupta AB Shrivastava GC On antibiotic properties of some fresh water algae Hydrobiologia 25 1 285 288 1965 sourceNot checked yet

  Not checked in the free run.

• cites [68] Chu CY Liao WR Huang R Lin LP Haemagglutinating and antibiotic activities of freshwater microalgae World J Microbiol Biotechnol 20 8 817 825 2004 sourceNot checked yet

  Not checked in the free run.

• cites [69] Santoyo S Rodríguez-Meizoso I Cifuentes A Green processes based on the extraction with pressurized fluids to obtain potent antimicrobials from Haematococcus pluvialis microalgae LWT Food Science Technol 42 7 1213 1218 2009 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Some algal chemicals accumulate within cells so they only act after the algae is damaged or ingested; in other species, toxins are secreted into the media by the algae to avoid negative interactions.

“Some of these chemicals accumulate within cells so they only act after the algae is damaged or ingested. In other species, toxins are secreted into the media by the algae to avoid negative interactions [70].”

• cites [70] Wolfe GV The chemical defense ecology of marine unicellular plankton: constraints, mechanisms, and impacts Biological Bull 198 2 225 2000 10786943 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Not all production strains can survive in extreme conditions, and there is still the possibility for an extremophile contamination to arise.

“Unfortunately, not all production strains can survive in extreme conditions, and there is still the possibility for an extremophile contamination to arise [62].”

• cites [62] Borowitzka MA Culturing microalgae in outdoor ponds Algal Culturing Techniques Andersen RA Academic Press NY, USA 218 2005 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Anaerobic digestion produces methane gas while keeping the majority of nutrients in a bacterial slurry that can be used for algal fertilizer.

“This bacterial process produces methane gas, while keeping the majority of the nutrients in a bacterial slurry that can be killed and the mix used for algal fertilizer.”

• cites [53] Sialve B Bernet N Bernard O Anaerobic digestion of microalgae as a necessary step to make microalgal biodiesel sustainable Biotechnol Advances 27 4 409 416 2009 19289163 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S2 (checkable).

Production estimates to date, primarily based on laboratory conditions in enclosed photobioreactors, will need to be recalculated for industrially scalable systems before they can be considered meaningful for biofuel production.

“production estimates to date, which are primarily based on laboratory conditions in enclosed photobioreactors, will need to be recalculated for industrially scalable systems before they can be considered meaningful for biofuel production”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Green algae have been used extensively in aquaculture, mainly for production of secondary metabolites such as β-carotene and astaxanthin.

“Green algae have been used extensively in aquaculture, mainly for the production of secondary metabolites, such as β-carotene and astaxanthin.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The US government-sponsored Aquatic Species Program provided initial characterization of a few hundred species from the 1970s through to the mid-1990s.

“The US government-sponsored Aquatic Species Program (ASP) provided initial characterization of a few hundred species, from the 1970s through to the mid-1990s [16].”

• cites [16] Sheehan J Dunahay T Benemann J Roessler P A Look Back at the U S Department of Energy's Aquatic Species Program – Biodiesel from Algae National Renewable Energy Laboratory CO, USA 328 1998 ▪ ▪ Outlines some of the early successes and challenges faced by groups currently addressing algae biofuels. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Botryococcus spp. produce triterpenoid botryoccenes, a potential fuel molecule that requires minimal refining.

“Botryococcus spp. that produce the triterpenoid botryoccenes, a potential fuel molecule that requires minimal refining [103].”

• cites [103] Metzger P Largeau C Botryococcus braunii: a rich source for hydrocarbons and related ether lipids Appl Microbiol Biotechnol 66 5 486 496 2005 15630516 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

A nitrogen-fixing cyanobacterium cocultured with a lipid-rich alga may form a synergistic relationship that allows total biomass density to expand above current estimates while minimizing the cost of adding nitrates.

“it is possible that a nitrogen-fixing cyanobacteria, cocultured with a lipid-rich algae, may form a synergistic relationship, allowing the total biomass density to expand above current estimates, while minimizing the cost of adding nitrates to the system”

• AssumptionRests on an assumption: Some cyanobacteria have the ability to fix nitrogen.

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

Algae are an extremely diverse group that contains many thousands of known species, and potentially hundreds of thousands.

“Algae are an extremely diverse group that contains many thousands of known species, and potentially hundreds of thousands.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Diatoms have more than 100,000 estimated species and are currently one of the most prolific primary marine producers.

“This diverse group, with more than 100,000 estimated species, is currently one of the most prolific primary marine producers.”

May combine two claims.

Algal growth in open pond systems has produced 0.06-0.231 g/l/day of biomass, and nearly 3 g/l/day in bioreactors.

“To date, algal growth in open pond systems have been able to produce 0.06–0.231 g/l/day of biomass [16,106,107] and nearly 3 g/l/day in bioreactors [108].”

• cites [16] Sheehan J Dunahay T Benemann J Roessler P A Look Back at the U S Department of Energy's Aquatic Species Program – Biodiesel from Algae National Renewable Energy Laboratory CO, USA 328 1998 ▪ ▪ Outlines some of the early successes and challenges faced by groups currently addressing algae biofuels. sourceNot checked yet

  Not checked in the free run.

• cites [106] Moheimani NR Borowitzka MA Limits to productivity of the alga Pleurochrysis carterae (haptophyta) grown in outdoor raceway ponds Biotechnol Bioeng 96 1 27 36 2007 16948168 sourceNot checked yet

  Not checked in the free run.

• cites [107] Blanco AM Moreno J Del Campo JA Rivas J Guerrero MG Outdoor cultivation of lutein-rich cells of Muriellopsis sp. in open ponds Appl Microbiol Biotechnol 73 6 1259 1266 2007 17033775 sourceNot checked yet

  Not checked in the free run.

• cites [108] Doucha J Livansky K Productivity, CO2/O2 exchange and hydraulics in outdoor open high density microalgal (Chlorella sp.) photobioreactors operated in a middle and southern european climate J Applied Phycology 18 6 811 826 2006 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

Nuclear transformation has been achieved in a number of other green algae species, such as members of the Chlorella and volvox genuses.

“while nuclear transformation has been achieved in a number of other green algae species, such as members of the Chlorella and volvox genuses [99,100].”

• cites [99] Lerche K Hallmann A Stable nuclear transformation of Gonium pectorale BMC Biotechnol 9 64 2009 19591675 PMC2720962 sourceNot checked yet

  Not checked in the free run.

• cites [100] Hawkins RL Nakamura M Expression of human growth hormone by the eukaryotic alga, Chlorella Curr Microbiol 38 6 335 341 1999 10341074 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

In silica-limited environments, diatoms have been found to accumulate lipids, but growth is reduced.

“In silica-limited environments, diatoms have been found to accumulate lipids; however, growth is reduced [88].”

• cites [88] Roessler PG Changes in the activities of various lipid and carbohydrate biosynthetic enzymes in the diatom Cyclotella cryptica in response to silicon deficiency Arch Biochem Biophys 267 2 521 528 1988 2905594 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Renewed interest in cyanobacteria as a biofuel source exists because some species can grow in extreme environments and because they have potential to be rapidly engineered.

“renewed interest in cyanobacteria as a potential biofuel source because of the ability of some species to grow in extreme environments, and the potential to rapidly engineer these species”

Probably not a substantial claim: may not pass S3 (load-bearing).

Cyanobacteria may be manipulated to secrete biofuel molecules.

“Cyanobacteria may also be able to be manipulated to secrete biofuel molecules.”

May combine two claims.

The diatom sexual lifecycle is poorly understood, making diatoms recalcitrant to traditional forward genetic exploration.

“Although other diploid species have become foundational model organisms (e.g. Arabidopsis, Drosophila and mice), the diatom sexual lifecycle is poorly understood, making them recalcitrant to traditional forward genetic exploration.”

Probably not a substantial claim: may not pass S3 (load-bearing).

It is believed that some oil deposits originated from diatom biomass.

“Furthermore, it is believed that some oil deposits originated from diatom biomass [87].”

• cites [87] Armbrust EV The life of diatoms in the World's oceans Nature 459 7244 185 192 2009 19444204 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

One reason for the focus on green algae is their shared common ancestry with vascular plants.

“One reason for this focus on green algae is their shared common ancestry with vascular plants.”

May combine two claims.

Whole-genome sequences have been released for Thalassiosira pseudonana and Phaeodactylum tricornutum, and stable nuclear transformation has been shown for these two species and a few other diatoms.

“Whole-genome sequences have been released for two species, Thalassiosira pseudonana and Phaeodactylum tricornutum, and recent work has shown stable nuclear transformation of these two species and a few other diatoms [91–94].”

• cites [91] Apt KE Kroth-Pancic PG Grossman AR Stable nuclear transformation of the diatom Phaeodactylum tricornutum Mol Gen Genet 252 5 572 579 1996 8914518Not checked yet

  Not checked in the free run.

• cites [92] Dunahay TG Jarvis EE Roessler PG Genetic transformation of the diatoms Cyclotella cryptica and Navicula saprophila J Phycol 31 6 1004 1012 1995 sourceNot checked yet

  Not checked in the free run.

• cites [93] Poulsen N Kroger N A new molecular tool for transgenic diatoms: control of mrna and protein biosynthesis by an inducible promoter-terminator cassette FEBS J 272 13 3413 3423 2005 15978046 sourceNot checked yet

  Not checked in the free run.

• cites [94] Falciatore A Casotti R Leblanc C Abrescia C Bowler C Transformation of nonselectable reporter genes in marine diatoms Mar Biotechnol (NY) 1 3 239 251 1999 10383998 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

These biomass numbers may improve through genetics and breeding, as has been accomplished in crop species.

“These numbers may improve through genetics and breeding, as has been accomplished in crop species”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The extant green algae consist of approximately 8000 species, found throughout freshwater and marine habitats, and a few species such as D. salina have been shown to survive in high or saturated saline solutions.

“The extant green algae consist of approximately 8000 species, and member species can be found throughout freshwater and marine habitats, and a few species (e.g., D. salina) have been shown to survive in high/saturated saline solutions.”

May combine two claims.

Optimizing the growth of algae in open ponds is a key component of reaching economic viability and remains a major challenge for the industry.

“Optimizing the growth of algae in open ponds is a key component of reaching economic viability, and remains a major challenge for the industry.”

May combine two claims.

The genomes of a number of green algae have been sequenced, and molecular tools are in place to transform both the chloroplast and nuclear genome of C. reinhardtii.

“The genomes of a number of green algae have been sequenced, and molecular tools are in place to transform both the chloroplast and nuclear genome of C. reinhardtii [96–98].”

• cites [96] Boynton JE Gillham NW Genetics and transformation of mitochondria in the green alga Chlamydomonas Methods Enzymol 264 279 296 1996 8965701 sourceNot checked yet

  Not checked in the free run.

• cites [97] Boynton JE Gillham NW Chloroplast transformation in Chlamydomonas Methods Enzymol 217 510 536 1993 8474349 sourceNot checked yet

  Not checked in the free run.

• cites [98] Kindle KL Schnell RA Fernandez E Lefebvre PA Stable nuclear transformation of Chlamydomonas using the Chlamydomonas gene for nitrate reductase J Cell Biol 109 6 Pt 1 2589 2601 1989 2592399 PMC2115893 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Lipid production in cyanobacteria may never reach that of eukaryotic algae.

“Although lipid production in cyanobacteria may never reach that of the eukaryotic algae”

May combine two claims.

A diatom species has been characterized that has endosymbiosed a nitrogen-fixing cyanobacterium, suggesting such relationships are beneficial to at least one of the two species.

“recently, a diatom species has been characterized, which has endosymbiosed a nitrogen-fixing cyanobacterium, suggesting that such relationships are indeed beneficial to at least one of the two species [104,105]”

• cites [104] Kneip C Voss C Lockhart PJ Maier UG The cyanobacterial endosymbiont of the unicellular algae rhopalodia gibba shows reductive genome evolution BMC Evol Biol 8 30 2008 18226230 PMC2246100 sourceNot checked yet

  Not checked in the free run.

• cites [105] Carpenter EJ Janson S Intracellular cyanobacterial symbionts in the marine diatom Climacodium frauenfeldianum (bacillariophyceae) J Phycol 36 3 540 544 2000 29544002 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S2 (checkable).

For true economic viability to be assessed, all aspects of algal physiology must be assessed under real-world conditions rather than theoretically achievable results under idealized conditions.

“in order for true economic viability to be assessed, and for algae to move forward from the bench to the pond, all aspects of algal physiology must be assessed under real-world conditions, rather than the theoretically achievable results under idealized conditions”

Probably not a substantial claim: may not pass S3 (load-bearing).

Green algae are the most heavily studied algal groups, primarily owing to the establishment of C. reinhardtii as an algal model organism.

“These green algae are the most heavily studied algal groups, primarily owing to the establishment of the chlorophyceae, C. reinhardtii, as an algal model organism.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The main producer of astaxanthin is the freshwater algae Haematococcus pluvialis, which can accumulate up to 4% of its dry weight as this pigment.

“The main producer of astaxanthin is the freshwater algae, Haematococcus pluvialis, which can accumulate up to 4% of its dry weight as this pigment [178]”

• cites [178] Eonseon J Lee CG Polle JEW Secondary carotenoid accumulation in Haematococcus (chlorophyceae): biosynthesis, regulation, and biotechnology J Microbiol Biotechnol 16 6 821 831 2006Not checked yet

  Not checked in the free run.

May combine two claims.

More than 20 different algae have now been transformed in either the nucleus or plastid, with all major classes having at least one report of foreign gene expression.

“more then 20 different algae have now been transformed in either the nucleus or plastid, with all major classes having at least one report of foreign gene expression”

May combine two claims.

Both the movement towards a hydrogen economy and the ability to engineer algal species to produce substantial amounts of hydrogen in a regulated manner have moved forward at a slower rate than once predicted.

“both the movement towards a hydrogen economy and the ability to engineer algal species to produce substantial amounts of hydrogen in a regulated manner have moved forward at a slower rate than once predicted”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

C. reinhardtii can and often does efficiently silence or downregulate nonrequired heterologous genes when expressed at high levels, perhaps as a defense against viral infection.

“C. reinhardtii can and often does efficiently silence or downregulate nonrequired heterologous genes when expressed at high levels, perhaps as a defense against viral infection.”

Probably not a substantial claim: may not pass S4 (based).

Traits from different species, when engineered into a single strain, may be sufficient to result in an economically viable production strain.

“These traits, when engineered into a single strain, may be sufficient to result in an economically viable production strain.”

May combine two claims.

Not all algae appear to show aggressive silencing of heterologous genes; the diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana have shown stable integration and expression of marker genes, including GFP.

“not all algae appear to show this aggressive silencing of heterologous genes. The diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana have both shown stable integration and expression of marker genes, including genes that do not necessarily impart a significant advantage (e.g., GFP)”

Probably not a substantial claim: may not pass S4 (based).

If infrastructure for algae fuel production is in place, expansion of facilities to include protein or other coproduct purification can be added at a fraction of total cost.

“If the infrastructure for algae fuel production is in place, expansion of these facilities to include protein or other coproduct purification can be added at a fraction of total cost.”

Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing), S4 (based).

'Omic fields will play a vital role when characterizing new species as researchers expand beyond classical model organisms.

“these 'omic fields will play a vital role when characterizing new species, as researchers expand beyond classical model organisms”

May combine two claims.

Directed and successful breeding of some species of microalgae is possible, but little is known about the sexual lifecycle of the vast majority of species.

“Directed and successful breeding of some species of microalgae is possible, but little is known about the sexual lifecycle of the vast majority of species.”

Probably not a substantial claim: may not pass S4 (based).

Many co-products will be coextracted with lipids using current strategies, decreasing their value as a coproduct.

“many of these co-products will be coextracted with the lipids using current strategies, decreasing their value as a coproduct”

May combine two claims.

Identification of an ideal, unmodified biofuel organism that fits established infrastructure and is economically viable is a possibility; however, a more likely scenario is identification of a variety of species each with one or a few desirable traits.

“Identification of an ideal, unmodified biofuel organism that fits into the established infrastructure for harvesting, extraction and purification, and is economically viable, is a possibility; however, a much more likely scenario is the identification of a variety of species that each have one or a few of these desirable traits.”

May combine two claims.

Chloroplast transformation occurs through homologous recombination rather than random integration and does not show gene silencing.

“chloroplast transformation occurs through homologous recombination rather than random integration and does not show gene silencing”

Probably not a substantial claim: may not pass S3 (load-bearing).

The primary traits of interest for algal biofuels are growth rates, growth density, lipid accumulation, resistance to predators and harvestability.

“The primary traits of interest are growth rates, growth density, lipid accumulation, resistance to predators and harvestability.”

May combine two claims.

β-carotene is produced by the marine algae D. salina, which can accumulate up to 14% of its dry weight as this pigment under stress conditions.

“β-carotene is produced by the marine algae D. salina, which can accumulate up to 14% of its dry weight as this pigment under stress conditions [175]”

• cites [175] Ye ZW Jiang JG Wu GH Biosynthesis and regulation of carotenoids in Dunaliella: progresses and prospects Biotechnol Advances 26 4 352 360 2008 18486405 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

Adding a reduced carbon source to algae cultures will increase the presence of unwanted pests, potentially making scale-up to volumes required for significant biofuel production problematic.

“adding a reduced carbon source to algae cultures will increase the presence of unwanted pests, potentially making the scale-up to the volumes required for significant biofuel production problematic”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The literature on algal strain improvements is minimal; however, extrapolation from successes in other systems may provide a blueprint for some improvements.

“In general, the literature on algal strain improvements is minimal; however, extrapolation from successes in other system may provide a blueprint for some of these improvements.”

Probably not a substantial claim: may not pass S2 (checkable).

These natural co-products have potential to provide a bridge while the economics of algal biofuels improve.

“These natural co-products have potential to provide a bridge while the economics of algal biofuels improve.”

May combine two claims.

Many of these traits are probably multigene traits, and species with sequenced genomes will offer significant advantages to those without.

“Many of these are probably multigene traits, and species with sequenced genomes will offer significant advantages to those without.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Modified quantitative trait loci strategies can be applied across species to identify orthologs that regulate similar traits in other species.

“there is potential to use modified quantitative trait loci (QTL) strategies, and apply these across species to identify orthologs that regulate similar traits in other species [113]”

• cites [113] Borevitz JO Maloof JN Lutes J Quantitative trait loci controlling light and hormone response in two accessions of Arabidopsis thaliana Genetics 160 2 683 696 2002 11861571 PMC1461994 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

There is potential to improve algae for biofuels by developing tools for selective breeding and using these to move traits that impact biofuels between closely related species or to improve specific strains of one species.

“There is potential to improve algae for biofuels by developing the tools for selective breeding and using these to move traits that have an impact on biofuels between closely related species, or to improve specific strains of one species.”

Probably not a substantial claim: may not pass S2 (checkable). May combine two claims.

Improving extraction techniques or dedicating a percentage of the algal crop to higher value products depending on demand may further close the economic gap between petroleum and algae biofuels.

“Improving extraction techniques or dedicating a percentage of the algal crop to these higher value products depending on demand (e.g., with terrestrial agriculture) may further close the economic gap between petroleum and algae biofuels (Table 2).”

May combine two claims.

The plastid genome of Chlamydomonas has been successfully transformed and has become a consistent method for heterologous protein expression.

“the plastid genome of Chlamydomonas has been successfully transformed and has become a consistent method for heterologous protein expression [117,118]”

• cites [117] Mayfield SP Manuell AL Chen S Chlamydomonas reinhardtii chloroplasts as protein factories Curr Opin Biotechnol 18 2 126 133 2007 17317144 sourceNot checked yet

  Not checked in the free run.

• cites [118] Purton S Tools and techniques for chloroplast transformation of Chlamydomonas Adv Exp Med Biol 616 34 45 2007 18161489 sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Algae produce hexose sugars, which can be fermented to produce ethanol, butanol and potentially longer chain hydrocarbons.

“algae produce hexose sugars, which can be fermented to produce ethanol, butanol and potentially longer chain hydrocarbons”

May combine two claims.

The enzymes involved in oil production may be manipulated to increase oil production, and the resulting oil can be used after transesterification as diesel or undergo carbon cracking to produce biogasoline or biojet fuels.

“The enzymes involved in oil production may be able to be manipulated, thus increasing the oil production of these species, and the resulting oil can be used after being transesterified as diesel or undergo carbon cracking to produce biogasoline or biojet fuels [161]”

• cites [161] Benson TJ Hernandez R White MG Heterogenous cracking of an unsaturated fatty acid and reaction intermediates on H+ZSM-5 catalyst Clean 36 8 652 656 2008Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based).

The economic impact from these strain improvements will allow for decreased operational costs.

“The economic impact from these improvements will allow for decrease operational costs.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Once extracted, the chemical profile of algal fatty acids is not expected to be substantially different from the chemical mixes currently found in sweet light crude oil.

“once extracted, its chemical profile is not expected to be substantially different from the chemical mixes currently found in sweet light crude oil”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Microalgae are amenable to many of the 'omic techniques developed for bacteria and single-celled eukaryotes, such as S. cerevisae.

“microalgae are amenable to many of the 'omic techniques developed for bacteria and single-celled eukaryotes, such as S. cerevisae”

Probably not a substantial claim: may not pass S3 (load-bearing).

DHA is an omega-3 fatty acid commercially produced by Crypthecodinium and Schizochytrium for infant formulas and aquaculture feeds.

“Docosahexaenoic acid (DHA) is an omega-3 fatty acid commercially produced by Crypthecodinium and Schizochytrium for infant formulas and aquaculture feeds.”

May combine two claims.

Arthrospira (Spirulina) has a 60-71% dry-weight protein content and is widely used as a food supplement for humans and animals.

“The cyanobacteria Arthrospira (i.e., Spirulina) has a 60–71% dry-weight protein content, and is widely used as a food supplement for humans, cattle, poultry, aquarium fish, ornamental birds and horses [168]”

• cites [168] Becker W Microalgae in human and animal nutrition Handbook of Microalgal Culture: Biotechnology & Applied Phycology Richmond A Blackwell Oxford, UK 312 351 2003 sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

An alternative coproduct strategy is to genetically engineer algae to produce higher value proteins, specifically industrial enzymes and animal feed supplements.

“An alternative coproduct strategy is to genetically engineer algae to produce higher value proteins, specifically industrial enzymes and animal feed supplements.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

Different methods of lipid extraction from hexane extraction to pressing extract lipids with different efficiencies and can yield different lipid profiles and returns.

“Different methods of lipid extraction from hexane extraction to pressing strategies extract lipids with different efficiencies and can yield different lipid profiles and returns.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Early on, diverse coproduct-producing strains will most likely be used rather than an optimal single strain.

“Early on, owing to the large market for fuels and companies establishing niches, it is most likely that diverse coproduct-producing strains will be used, rather than an optimal single strain.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Other strain-specific improvements being considered include crop protection, salt tolerance, growth at high pH, improved nutrient utilization and traits for more efficient harvesting such as flocculation.

“other strain-specific improvements are being considered, including crop protection, salt tolerance, growth at high pH, improved nutrient utilization and traits that lead to more efficient harvesting, such as flocculation”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

The ability to secrete lipids may improve efficiency in harvesting and nutrient utilization, as well as provide some protection from contamination.

“The ability to secrete lipids may improve efficiency in harvesting and nutrient utilization, as well as provide some protection from contamination.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based). May combine two claims.

Flocculation will allow improved harvesting by making it easier to concentrate algae, decreasing the cost of water extraction.

“flocculation will allow improved harvesting by making it easier to concentrate algae, decreasing the cost of water extraction”

May combine two claims.

Fuel production from algae can be cost competitive and widely scalable and deployable in the next 7-10 years, but only if understanding of algae and ability to engineer them continue to expand.

“Even given these uncertainties, we believe that fuel production from algae can be cost competitive and widely scalable and deployable in the next 7–10 years”

• Also stated in: § Future perspective · plain

  “but only if we continue to expand our understanding of these amazing organisms as we expand our ability to engineer them for the specific task of developing a new energy industry.”

Probably not a substantial claim: may not pass S4 (based).

Subsequent genetic engineering and breeding will be required to bring these strains to economic viability.

“and subsequent genetic engineering and breeding will be required to bring these strains to economic viability.”

Probably not a substantial claim: may not pass S2 (checkable), S4 (based).

The most likely scenario is that bioprospecting will not identify algal species that are cost competitive with petroleum.

“Despite the potential of this strategy, the most likely scenario is that bioprospecting will not identify species that are cost competitive with petroleum”

May combine two claims.

Many industrial enzymes can be produced with values ranging between $25 and $50 per kg, and large markets currently exist for them.

“there are many industrial enzymes that can be produced, with values ranging between $25 and $50 per kg, and for which large markets currently exist [25]”

• cites [25] Rosenberg JN Oyler GA Wilkinson L Betenbaugh MJ A green light for engineered algae: redirecting metabolism to fuel a biotechnology revolution Curr Opin Biotechnol 19 5 430 436 2008 18725295 sourceNot checked yet

  Not checked in the free run.