Methods of oil production in microorganisms

Abstract

Provided herein are methods for producing one or more polyunsaturated fatty acids. The methods include the steps of providing a microorganism capable of producing polyunsaturated fatty acids, providing a medium comprising a high concentration of one or more carbon sources, low pH, or both, and culturing the microorganism in the medium under sufficient conditions to produce the one or more polyunsaturated fatty acids. Also provided are methods of culturing one or more microorganisms. The methods include culturing the microorganisms in a medium comprising a first amount of one or more carbon sources at a first concentration level, monitoring a carbon source concentration until the carbon source concentration is reduced below the first concentration level, and adding to the medium a second amount of one or more carbon sources to increase the carbon source concentration to a second concentration level.

What is claimed is: 
     
         1 . A method for producing one or more polyunsaturated fatty acids, the method comprising:
 (a) providing a microorganism capable of producing polyunsaturated fatty acids;   (b) providing a medium comprising a high concentration of one or more carbon sources, low pH, or both; and   (c) culturing the microorganism in the medium under sufficient conditions to produce the one or more polyunsaturated fatty acids.   
     
     
         2 . The method of  claim 1 , wherein the medium has a low pH. 
     
     
         3 . The method of  claim 1 , wherein the medium has a high concentration of one or more carbon sources. 
     
     
         4 . The method of  claim 1 , wherein the medium has a low pH and a high concentration of one or more carbon sources. 
     
     
         5 . The method of  claim 1 , wherein the microorganism is ONC-T18. 
     
     
         6 . The method of  claim 1 , wherein the concentration of the one or more carbon sources is 200 to 300 g/L. 
     
     
         7 . The method of  claim 1 , wherein the carbon source is selected from the group consisting of fatty acids, lipids, glycerols, triglycerols, carbohydrates, polyols, and amino sugars. 
     
     
         8 . The method of  claim 1 , wherein the carbon source is glucose. 
     
     
         9 . The method of  claim 1 , wherein the pH of the medium is 2 to 4.5. 
     
     
         10 . The method of  claim 9 , wherein the pH of the medium is 3 to 3.5. 
     
     
         11 . The method of  claim 1 , further comprising isolating the polyunsaturated fatty acids, wherein the medium is not sterilized prior to isolation of the polyunsaturated fatty acids. 
     
     
         12 . The method of  claim 11 , wherein sterilization comprises an increase in temperature. 
     
     
         13 . A method of reducing contamination of a non-sterile culture comprising one or more microorganisms, the method comprising culturing the microorganisms (i) in the presence of a high concentration of one or more carbon sources, (ii) under conditions of low pH, or (iii) a combination thereof, wherein the culturing reduces contamination of the non-sterile culture comprising the microorganisms. 
     
     
         14 . The method of  claim 13 , wherein the method comprises culturing the microorganisms in an open vessel. 
     
     
         15 . The method of  claim 13 , wherein the culturing comprises culturing the microorganisms in the presence of a high concentration of one or more carbon sources. 
     
     
         16 . The method of  claim 13 , wherein the culturing comprises culturing the microorganisms under conditions of low pH. 
     
     
         17 . The method of  claim 13 , wherein the culturing comprises culturing the microorganisms in the presence of a high concentration of one or more carbon sources and under conditions of low pH. 
     
     
         18 . The method of  claim 13 , wherein the microorganism is ONC-T18. 
     
     
         19 . The method of  claim 13 , wherein the concentration of the one or more carbon sources is 200 to 300 g/L. 
     
     
         20 . The method of  claim 13 , wherein the carbon source is selected from the group consisting of fatty acids, lipids, glycerols, triglycerols, carbohydrates, polyols, and amino sugars. 
     
     
         21 . The method of  claim 13 , wherein the carbon source is glucose. 
     
     
         22 . The method of  claim 13 , wherein the pH of the medium is 2 to 4.5. 
     
     
         23 . The method of  claim 22 , wherein the pH of the medium is 3 to 3.5. 
     
     
         24 . A method of culturing one or more microorganisms comprising:
 (a) culturing the microorganisms in a medium comprising a first amount of one or more carbon sources at a first concentration level, wherein the first concentration level of the one or more carbon sources is greater than 200 g/L;   (b) monitoring a carbon source concentration until the carbon source concentration is reduced below the first concentration level; and   (c) adding to the medium a second amount of one or more carbon sources to increase the carbon source concentration to a second concentration level, wherein the second concentration level of the one or more carbon sources is greater than 200 g/L.   
     
     
         25 . The method of  claim 24 , wherein the second amount of the one or more carbon sources is added to the medium when the carbon source concentration level is reduced to 0 to 20 g/L. 
     
     
         26 . The method of  claim 24 , further comprising, after addition of the second amount of the one or more carbon sources, (a) culturing the microorganisms until the carbon source concentration of the one or more carbon sources is reduced below the second concentration level and (b) adding to the medium a third amount of one or more carbon sources to increase the carbon source concentration to a third concentration level. 
     
     
         27 . The method of  claim 26 , wherein the third concentration level of the one or more carbon sources is greater than 200 g/L. 
     
     
         28 . The method of  claim 26 , wherein the third amount of the one or more carbon sources is added to the medium when the carbon source concentration is reduced to 0 to 20 g/L. 
     
     
         29 . The method of  claim 26 , further comprising, after addition of the third amount of the one or more carbon sources, (a) culturing the microorganisms until the carbon source concentration of the one or more carbon sources is reduced below the third concentration level and (b) adding to the medium a fourth amount of one or more carbon sources to increase the carbon source concentration to a fourth concentration level. 
     
     
         30 . The method of  claim 29 , wherein the fourth concentration level of the one or more carbon sources is greater than 200 g/L. 
     
     
         31 . The method of  claim 29 , wherein the fourth amount of the one or more carbon sources is added to the medium when the carbon source concentration is reduced to 0 to 20 g/L. 
     
     
         32 . The method of  claim 24 , wherein the methods further comprise monitoring the carbon source concentration one or more times. 
     
     
         33 . The method of  claim 32 , wherein the monitoring comprises measuring dissolved oxygen levels. 
     
     
         34 . The method of  claim 32 , wherein monitoring comprises obtaining a sample of the medium and determining the carbon source concentration in the sample. 
     
     
         35 . The method of  claim 24 , wherein the microorganism is ONC-T18. 
     
     
         36 . The method of  claim 24 , wherein the microorganisms are capable of producing one or more polyunsaturated fatty acids. 
     
     
         37 . The method of  claim 36 , wherein the method further comprises isolating the polyunsaturated fatty acids produced by the microorganisms. 
     
     
         38 . The method of  claim 34 , wherein the carbon source is selected from the group consisting of fatty acids, lipids, glycerols, triglycerols, carbohydrates, polyols, and amino sugars. 
     
     
         39 . The method of  claim 34 , wherein the carbon source is glucose. 
     
     
         40 . The method of  claim 29 , wherein the one or more carbon sources in the first, second, third, and fourth amounts are the same.