Methods and Compositions for Regulating Sporulation
Methods and compositions are provided for producing fuel utilizing various strains of Clostridium phytofermentans with reduced sporulation activity. In some embodiments, the activity of a gene associated with sporulation is reduced. In some embodiments, a fuel producing strain of C. phytofermentans with reduced sporulation activity is provided.
1 . A method for producing a compound, the method comprising:
incubating a culture medium with a Clostridium phytofermentans cell, wherein said cell has altered activity of at least one gene associated with sporulation as compared to the activity of the gene in a wild-type C. phytofermentans strain, and wherein said altered activity enhances production of said compound; and
isolating the compound from the culture medium.
2 . The method of claim 1 , wherein said compound is ethanol.
3 . The method of claim 1 , wherein said compound comprises at least one solvent selected from the group consisting of propanol, proprionate, acetate, lactate, and formate.
4 . The method of claim 1 , wherein said gene associated with sporulation is a gene upregulated by Spo0A.
5 . The method of claim 1 , wherein said gene associated with sporulation is selected from the group consisting of SpoIIAA, SpoIIAB, SigF, SpoIIE, SpoIIGA, SigG, and SigE.
6 . The method of claim 1 , wherein said gene associated with sporulation is SpoIIE.
7 . The method of claim 6 , wherein said altered activity is reduced activity of said gene.
8 . The method of claim 1 , wherein said altered activity is obtained by introducing into the cell an oligonucleotide that targets the gene.
9 . The method of claim 8 , wherein said oligonucleotide is introduced into the cell by expression from a plasmid that has been introduced into the cell.
10 . The method of claim 1 , wherein said altered activity is obtained by deleting or mutating at least a portion of the gene associated with sporulation.
11 . The method of claim 1 , wherein the culture medium comprises a biomass material.
12 . The method of claim 11 , wherein the biomass material comprises a plant polysaccharide.
13 . The method of claim 11 , wherein the biomass material comprises a lignocellulosic material.
14 . An isolated Clostridium phytofermentans cell comprising altered sporulation activity in comparison to a wild-type strain of C. phytofermentans.
15 . The cell of claim 14 , comprising an alteration of at least one gene associated with sporulation.
16 . The cell of claim 14 , wherein said gene is selected from the group consisting of SpoIIAA, SpoIIAB, SigF, SpoIIE, SpoIIGA, SigG, and SigE.
17 . The cell of claim 15 , wherein said gene is upregulated by Spo0A.
18 . The cell of claim 15 , wherein said gene is SpoIIE.
19 . The cell of claim 15 , wherein said alteration comprises disruption of gene activity by an oligonucleotide.
20 . The cell of claim 15 , wherein said alteration comprises at least a partial deletion or mutation.
21 . The cell of claim 19 , further comprising an expression plasmid encoding said oligonucleotide.
22 . A method for reducing sporulation in a Clostridium phytofermentans cell, the method comprising reducing the activity of a gene associated with sporulation.
23 . The method of claim 22 , wherein reducing comprises disrupting the gene.
24 . The method of claim 22 , wherein said reducing the activity comprises providing an oligonucleotide to the strain.
25 . The method of claim 24 , wherein the oligonucleotide is provided by an expression plasmid.
26 . The method of claim 22 , wherein reducing the activity comprises deleting or mutating at least a portion of the gene.
27 . A fuel producing mixture comprising:
an isolated Clostridium phytofermentans microorganism comprising an alteration of at least one gene associated with sporulation; and
biomass.
28 . The mixture of claim 27 , wherein the gene is selected from the group consisting of SpoIIAA, SpoIIAB, SigF, SpoIIE, SpoIIGA, SigG, and SigE.
29 . The mixture of claim 27 , wherein the gene is upregulated by Spo0A.
30 . The mixture of claim 27 , wherein the gene is SpoIIE.
31 . The mixture of claim 27 , further comprising a second microorganism.
32 . The mixture of claim 31 , wherein the second microorganism is a yeast, fungus, or bacterium.