Heterotrophic production methods for microbial biomass and bioproducts 机翻标题: 暂无翻译,请尝试点击翻译按钮。

公开号/公开日
US2018002711 A1 2018-01-04 [US20180002711] / 2018-01-04
申请号/申请日
2017US-15640246 / 2017-06-30
发明人
SCHURR ROBERT J;KUEHNLE ADELHEID R;
申请人
KUEHNLE AGROSYSTEMS;
主分类号
IPC分类号
A01N-065/03A23K-020/10A23K-050/80C12N-001/12C12N-015/82C12P-003/00C12P-005/02C12P-007/26C12P-009/00G01N-033/50
摘要
(US20180002711) The invention pertains to a method for synthesizing a product of interest by culturing a microalgal cell producing the product of interest in the dark in a culture medium comprising an organic acid as a fixed carbon source, wherein the microalgal cell is a facultative heterotroph.  The product of interest can be a microalgal biomass, a pigment, terpene, recombinant molecule, biogas, or a precursor thereof.  In an embodiment, the culture medium comprises urea as a primary source of nitrogen.  In one embodiment, the microalgal cell belongs to the order Chlamydomonadales.  A method of identifying and isolating a microalgal cell having a preferred characteristic that is suitable for synthesis of a product of interest is also provided, the method comprising identifying and isolating a non-mutagenized or recombinant microalgal cell from a microalgal culture using a fluorescence activated cell sorting technique and/or a phototaxic response.
机翻摘要
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地址
代理人
代理机构
;
优先权号
2016US-62356896 2016-06-30 2017US-15640246 2017-06-30
主权利要求
(US20180002711) What is claimed is: 1.  A method for synthesizing a product of interest, the method comprising: providing a culture medium comprising an organic acid as a fixed carbon source;   providing a microalgal cell that produces the product of interest, wherein the microalgal cell is a facultative heterotroph;   culturing the microalgal cell in the culture medium in the dark to produce a microalgal culture from the microalgal cell;   isolating the microalgal cells from the microalgal culture before the cells in the microalgal culture undergo cell differentiation; and   purifying the product of interest from the microalgal cells. 2. The method of claim 1, wherein the microalgal cell is a facultative heterotroph that is rendered an obligate heterotroph. 3. The method of claim 1, wherein the product of interest is a microalgal biomass comprising the microalgal cells. 4. The method of claim 1, wherein the product of interest is a pigment, terpene, recombinant molecule, biogas, or a precursor thereof. 5. The method of claim 4, wherein the pigment is a carotenoid, isoprenoid, or precursor thereof. 6. The method of claim 4, wherein the pigment or a precursor thereof is astaxanthin, lutein, lycopene, zeaxanthin, canthaxanthin, carotene, phytofluene, phytoene, or any combination thereof. 7. The method of claim 4, wherein the terpene or a precursor thereof is pinene, limonene, or geranylgeranyl pyrophosphate or any combination thereof. 8. The method of claim 4, wherein the recombinant molecule is a dsRNA or heterologous protein. 9. The method of claim 5, wherein the microalgal culture produces the pigment at a specific productivity rate, qp, of at least 0.063 mg/L-hour. 10. The method of claim 1, wherein the step of culturing is conducted for a period of less than one week. 11. The method of claim 1, wherein the heterotrophic growth and synthesis of the product of interest occurs during the step of culturing, and wherein the step of culturing is performed under a fed-batch fermentation. 12. The method of claim 1, wherein the synthesis of the product of interest occurs under nutrient replete or deplete conditions. 13. The method of claim 1, wherein the organic acid is acetic, citric, fumaric, glycolic, lactic, malic, proprionic, pyruvic, succinic, glucuronic, galacturonic, uronic, chlorogenic, or lignocellulosic acid. 14. The method of claim 1, wherein the culture medium comprises urea as a primary source of nitrogen. 15. The method of claim 1, wherein the method further comprises one or more steps of drying, grinding, lysing, and extracting the microalgal cell. 16. The method of claim 1, wherein the microalgal cell belongs to the order Chlamydomonadales. 17. The method of claim 16, wherein the microalgal cell is a Haematococcus spp., Chlamydomonas spp., Chloromonas spp., Dunaliella spp., or Chlamydocapsa spp. 18. The method of claim 17, wherein the microalgal cell is replaced by a microbial cell type that preferentially metabolizes organic acid and urea. 19. The method of claim 1, wherein the microalgal cell is co-cultivated with a second cell. 20. The method of claim 19, wherein the second cell uses a different fixed carbon source to support its growth compared to the organic acid which supports the growth of the microalgal cell. 21. The method of claim 20, wherein the second cell consumes ammonium as a nitrogen source. 22. The method of claim 21, wherein the second cell type belongs to Scenedesmus spp., Chlorella spp., Monoraphidium spp., Rhodotorula spp., a diatom, a thraustochytrid, or a thraustochytrid-like microorganism. 23. The method of claim 19, wherein the microalgal cell belongs to a Haematococcus spp. and the second cell belongs to a Chlamydomonas spp. 24. A facultative heterotrophic microalgal cell from the order Chlamydomonadales, wherein said cell produces a product when grown in the dark in a medium containing only organic acid as a primary source of carbon and before cell differentiation occurs; and wherein said cell is a recombinant cell expressing nucleic acids for one or more RNA or protein for the production of pinene, limonene, astaxanthin, phytoene, lycopene, geranylgeranyl pyrophosphate, biohydrogen, or thaumatin; or wherein said cell is a recombinant cellexpressing a dsRNA molecule. 25. A facultative heterotrophic microalgal cell from the order Chlamydomonadales, wherein said cell produces a product when grown in the dark in a medium containing only organic acid as a primary source of carbon and before cell differentiation occurs; and wherein said cell is a mutant cell with altered production of isoprenoids or is a flagellum mutant. 26. The facultative heterotrophic cell of claim 25, wherein the cell has altered isoprenoid or chlorophyll production relative to the original strain. 27. The facultative heterotrophic cell of claim 25, wherein the cell is directly selected for altered production of carotenoid using fluorescence activated cell sorting. 28. The facultative heterotrophic cell of claim 25, wherein the cell is indirectly selected for altered production of non-carotenoid terpene using fluorescence activated cell sorting. 29. A method of identifying and isolating a microalgal cell that is suitable for synthesis of a product of interest, the method comprising: a. culturing a microalgal strain under at least partially heterotrophic conditions to produce microalgal cells;   b. identifying a non-mutagenized microalgal cell having a preferred characteristic, when grown in heterotrophic conditions, relative to the microalgal strain from which the microalgal cells are produced, where the step of identifying is performed using a fluorescence activated cell sorting technique and/or a phototaxic response; and   c. isolating the non-mutagenized microalgal cell having the preferred characteristic. 30. The method of claim 29, wherein the step of culturing is performed under mixotrophic conditions at least for a portion of the culturing step. 31. The method of claim 29, wherein the preferred characteristic is an increased synthesis of a product of interest by the microalgal cell. 32. The method of claim 29, wherein the preferred characteristic is the absence of flagella in the microalgal cell. 33. The method of claim 29, wherein the non-mutagenized microalgal cell is a recombinant cell that is indirectly selected for altered production of non-carotenoid terpene or directly selected for altered production of carotenoid using fluorescence activated cell sorting. 34. A colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient obtained from a heterotrophic microalgal biomass of Chlamydomonadales having a composition comprising more than 0.7% dry weight carotenoids, more than 40% dry weight protein, less than 10% dry weight ash, and a mixture of two or more carotenoids. 35. The colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient according to claim 34 that is a carotenoid-containing lysate or extract. 36. The colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient according to claim 34 that additionally contains microalgal fatty acids. 37. The method of claim 12, wherein the nutrient replete or deplete conditions include one or more of sulfate, phosphate or urea depletion, or ammonium accumulation in excess of 2.5 mM; 38. The method of claim 12, wherein the nutrient replete or deplete condition are combined with one or more extrinsic factors selected from an addition of osmoticum greater than 2.6 g/L, addition of 45 mM NaCl, addition of lactic acid in excess of 3 g/L, an increase in temperature 2 degrees above the growth temperature; or any combination thereof. 39. The method of claim 1, wherein the microalgal culture is grown under mixing of 100 rpm or greater. 40. The microalgal biomass of claim 3 is used in salmonid feed. 41. The microalgal biomass of claim 3 that is used as a larvicide. 42. The microalgal biomass of claim 3 that is used as a flavor or fragrance modifier. 43. The colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient according to claim 34, whose profile varies according to the microalgal culture conditions. 44. The colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient according to claim 34, in which one carotenoid comprises greater than 50% of the total carotenoids. 45. The colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient according to claim 34, in which one carotenoid comprises greater than 85% of the total carotenoids. 46. The colorant, feed additive, human nutrient supplement, or personal care or cosmetics ingredient according to claim 34, in which one carotenoid comprises greater than 98% of the total carotenoids. 47. The one carotenoid of claim 44, in which the carotenoid is astaxanthin. 48. The method of claim 8, wherein the dsRNA is produced at a specific productivity rate, qp, of at least 0.04 mg/L-hour. 49. The method of claim 8, wherein the heterologous protein is produced at a specific productivity rate, qp, of at least 0.08 mg/L-hour. 50. The microalgal biomass of claim 3 that is used in a seed train. 51. The method of claim 1, wherein the microalgal culture has a specific growth rate of at least 0.24/day. 52. The method of claim 51, wherein the microalgal culture has a specific growth rate of at least 0.77/day. 53. The method of claim 51, wherein the microalgal culture has a specific growth rate of at least 1.0/day. 54. The method of claim 51 wherein the specific growth rate is over a 96-hour period or longer. 55. The method of claim 52, wherein the specific growth rate is over a 96-hour period or longer. 56. The method of claim 53, wherein the specific growth rate is over a 96-hour period or longer. 57. The method of claim 9, wherein the qp is at least 1 mg/L-hour. 58. The method of claim 9, wherein the qp is at least 3.75 mg/L-hour. 59. The method of claim 10, wherein the step of culturing is conducted for a period of about 4 days to about one week. 60. The method of claim 54, wherein the microalgal culture with the specific growth rate over a 96-hour period or longer proceeds through cellular differentiation in the absence of light.
法律状态
(US20180002711) LEGAL DETAILS FOR US2018002711  Actual or expected expiration date=2037-06-30    Legal state=ALIVE    Status=PENDING     Event publication date=2017-06-30  Event code=US/APP  Event indicator=Pos  Event type=Examination events  Application details  Application country=US US15640246  Application date=2017-06-30  Standardized application number=2017US-15640246     Event publication date=2017-06-30  Event code=US/ART  Event type=Administrative notifications  USPTO Art Group  ART=1653     Event publication date=2017-06-30  Event code=US/DK  Event type=Examination events  Attorney Docket Number Docket Nbr: KAS.107X    Event publication date=2017-06-30  Event code=US/CUST  Event type=Examination events  Attorney/Agent Customer Number Customer Nbr: 23557    Event publication date=2017-06-30  Event code=US/SMALL  Event type=Administrative notifications  Appl Has Filed a Verified Statement of Micro to Small Entity Status Business Entity Status: SMALL    Event publication date=2017-06-30  Event code=US/AIA  Event type=Administrative notifications  First Inventor File Indicated:  AIA=Yes     Event publication date=2017-06-30  Event code=US/ENT  Event type=Administrative notifications  Entity Status Set to Undiscounted    Event publication date=2017-07-11  Event code=US/SMALL  Event type=Administrative notifications  Appl Has Filed a Verified Statement of Micro to Small Entity Status    Event publication date=2017-07-13  Event code=US/APPFILEREC  Event type=Administrative notifications  Event type=OAO  Filing Receipt    Event publication date=2017-09-13  Event code=US/PTARDY  Event indicator=Pos  Event type=Examination events  Patent Term Adjustment - Ready for Examination    Event publication date=2017-09-20  Event code=US/POAC  Event type=Change of name or address  Change in Power of Attorney (May Include Associate POA)    Event publication date=2017-09-20  Event code=US/RECEIPTUP  Event type=Administrative notifications  Event type=OAO  Filing Receipt - Updated    Event publication date=2017-11-30  Event code=US/IDS  Event type=Examination events  Event type=OAI  Information Disclosure Statement Filed    Event publication date=2018-01-04  Event code=US/A1  Event type=Examination events  Application published  Publication country=US  Publication number=US2018002711  Publication stage Code=A1  Publication date=2018-01-04  Standardized publication number=US20180002711     Event publication date=2018-01-04  Event code=US/PGPUBN  Event indicator=Pos  Event type=Examination events  Event type=OAO  PG-Pub Issue Notification    Event publication date=2018-01-29  Event code=US/IDS  Event type=Examination events  Event type=OAI  Information Disclosure Statement Filed    Event publication date=2018-03-12  Event code=US/DKNC  Event indicator=Pos  Event type=Examination events  Docketed New Case - Ready for Examination    Event publication date=2018-03-12  Event code=US/DOCK  Event indicator=Pos  Event type=Examination events  Case Docketed to Examiner
专利类型码
A1
国别省市代码
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