Genetically Modified Photoautotrophic Ethanol Producing Host Cells, Method For Producing The Host Cells, Constructs For The Transformation Of The Host Cells, Method For Testing A Photoautotrophic Strain For A Desired Growth Property And Method Of Producing Ethanol Using The Host Cells
The invention provides genetically modified photoautotrophic ethanol producing host cells. Ethanol production is controlled by an inducible promoter. Promoters utilized may be endogenous or heterologous, and genetic modification may be obtained by extrachromosomal means or chromosomal insertion.
1 . A recombinant cyanobacterial host cell comprising an exogenous polynucleotide sequence that binds to a polynucleotide sequence encoding a component of pyruvate dehydrogenase (EC number 1.2.4.1).
2 . The recombinant cyanobacterial host cell of claim 1 further comprising exogenous genes selected from the group consisting of adh and pdc.
3 . A recombinant cyanobacterial host cell comprising a vector comprising an exogenous polynucleotide sequence whose transcription is operably linked to a promoter wherein a transcriptional product of said exogenous polynucleotide sequence binds to a mRNA polynucleotide produced from transcription of an endogenous polynucleotide sequence encoding pyruvate dehydrogenase (EC number 1.2.4.1), and wherein said transcriptional product of said exogenous polynucleotide sequence binds to said mRNA polynucleotide and reduces translation of said mRNA polynucleotide in comparison to the translation of said mRNA polynucleotide wherein said exogenous polynucleotide sequence is not transcribed.
4 . The recombinant cyanobacterial host cell of claim 3 wherein said promoter is selected from the group consisting of ntcA, nblA, isiA, petJ, petE, ggpS, psbA2, psaA, sigB, IrtA, htpG, nirA, hspA, clpB1, hliB, and crhC.
5 . The recombinant cyanobacterial host cell of claim 3 wherein said host cell is a member of the genus Synechocystis.
6 . The recombinant cyanobacterial host cell of claim 3 wherein said exogenous polynucleotide sequence has an identity of greater than 70% to a polynucleotide sequence consisting of the sequence of nucleotides 5 through 225 of SEQ ID NO: 24.
7 . The recombinant cyanobacterial host cell of claim 3 wherein said exogenous polynucleotide sequence has an identity of greater than 70% to a polynucleotide sequence consisting of the sequence of nucleotides 8 through 90 of SEQ ID NO: 24.
8 . The recombinant cyanobacterial host cell of claim 3 further comprising an exogenous adh gene and an exogenous pdc gene wherein transcription of said adh gene is operably linked to a promoter and said pdc gene is operably linked to a promoter.
9 . The recombinant cyanobacterial host cell of claim 8 wherein said promoter of said exogenous adh gene and said promoter of said exogenous pdc gene are the same promoter.
10 . The recombinant cyanobacterial host cell of claim 9 wherein said promoter is selected from the group consisting of ntcA, nblA, isiA, petJ, petE, ggpS, psbA2, psaA, sigB. IrtA, htpG, nirA, hspA, clpB1, hliB, crhC and rbcLS.
11 . The recombinant cyanobacterial host cell of claim 8 wherein said promoter of said exogenous adh gene and said promoter of said exogenous pdc gene are different promoters.
12 . The recombinant cyanobacterial host cell of claim 11 wherein said promoters are selected from the group consisting of ntcA nblA, isiA pet, petE, ggpS, psbA2, psaA, sigB, IrtA, htpG, nirA, hspA, clpB1, hliB, crhC and rbcLS.
13 . A recombinant cyanobacterial host cell comprising a disruption of an endogenous polynucleotide sequence encoding pyruvate dehydrogenase (EC number 1.2.4.1), and further comprising a vector comprising an exogenous polynucleotide sequence encoding pyruvate dehydrogenase (EC number 1.2.4.1), wherein transcription of said exogenous polynucleotide sequence encoding pyruvate dehydrogenase (EC number 1.2.4.1) is operably linked to an inducible promoter.
14 . The recombinant cyanobacterial host cell of claim 13 wherein said inducible promoter is selected from the group consisting of ntcA, nblA, isiA, petJ, petE, ggpS, psbA2, psaA, sigB, IrtA, htpG, nirA, hspA, clpB1 hliB, and crhC.
15 . The recombinant cyanobacterial host cell of claim 13 wherein said host cell is a member of the genus Synechocystis.
16 . The recombinant cyanobacterial host cell of claim 13 further comprising an exogenous adh gene and an exogenous pdc gene wherein transcription of said adh gene is operably linked to a promoter and said pdc gene is operably linked to a promoter.
17 . The recombinant cyanobacterial host cell of claim 16 wherein said promoter of said exogenous adh gene and said promoter of said exogenous pdc gene are the same promoter.
18 . The recombinant cyanobacterial host cell of claim 17 wherein said promoter is selected from the group consisting of ntcA, nblA, isiA, pet, petE, ggpS, psbA2, psaA sigB, IrtA, htpG, nirA, hspA, clpB1 hliB, crhC and rbcLS.
19 . The recombinant cyanobacterial host cell of claim 16 wherein said promoter of said exogenous adh gene and said promoter of said exogenous pdc gene are different promoters.
20 . The recombinant cyanobacterial host cell of claim 19 wherein said promoters are selected from the group consisting of ntcA, nblA, isiA, petJ, petE, ggpS, psbA2, psaA, sigB, IrtA, htpG, nirA, hspA, clpB1, hliB, crhC and rbcLS.
21 . The recombinant cyanobacterial host cell of claim 13 wherein said exogenous polynucleotide sequence encodes for an enzyme that has an amino acid sequence that has an identity of greater than 78% to an amino acid sequence of pyruvate dehydrogenase from Synechocystis PCC 6803, accession number NP — 440765.