Down regulation of auxin response factor NbTF7 to increase nicotine in a plant
View Patent ↗Plant metabolism and alkaloid levels can be regulated by transcription factors that regulate the nicotinic alkaloid biosynthetic pathway. In one embodiment, the disclosure provides a transcription factor that negatively regulates alkaloid biosynthesis, such as nicotine biosynthesis. In particular, the present disclosure provides methods for the inhibition of Nicotiana benthamiana auxin response factor (NbTF7) to increase alkaloid biosynthesis in plants.
1. A method for increasing nicotine in a Nicotiana benthamiana or tabacum plant, comprising
introducing into a Nicotiana benthamiana or tabacum plant cell a vector comprising at least 21 consecutive nucleotides of a nucleic acid molecule comprising a nucleotide sequence selected from the group consisting of:
(i) the nucleotide sequence set forth in SEQ ID NO: 14 or SEQ ID NO: 15;
(ii) a nucleotide sequence that encodes a polypeptide having the amino acid sequence set forth in SEQ ID NO: 16; and
(iii) a nucleotide sequence that is at least 90% identical to the nucleotide sequences of (i) or (ii); and
wherein the consecutive nucleotides are in sense orientation, antisense orientation, or both.
2. The method of claim 1 , further comprising overexpressing within the Nicotiana plant at least one of berberine bridge enzyme-like protein a (NBB1), isoflavone reductase homolog A622 (A622), quinolate phospho-ribosyltransferase (QPT), putrescine-N-methyltransferase (PMT), and N-methylputrescine oxidase (MPO).
3. The method of claim 1 , further comprising overexpressing at least one transcription factor that positively regulates nicotine biosynthesis.
4. The method of claim 1 , further comprising suppressing the expression of at least one transcription factor that negatively regulates nicotine biosynthesis.
5. The method of claim 1 , wherein the plant is a Nicotiana tabacum plant.
6. A transgenic Nicotiana benthamiana or tabacum plant produced by the method of claim 1 , wherein the plant has an increased nicotine content, as compared to a control plant grown under similar conditions.
7. A product comprising the transgenic Nicotiana benthamiana or tabacum plant of claim 6 or portions thereof, wherein the product has an increased level of nicotine as compared to a product produced from a control plant.
8. The product of claim 7 , wherein the product is selected from the group consisting of a cigarette, cigarette tobacco, cigar tobacco, a cigar, pipe tobacco, chewing tobacco, snuff, snus, lozenges, and reconstituted tobacco.
9. Seeds from the transgenic Nicotiana benthamiana or tabacum plant of claim 6 , wherein the seeds comprise the nucleic acid molecule.
10. The method of claim 1 , wherein the nucleotide sequence comprises the nucleotide sequence set forth in SEQ ID NO: 14.
11. The method of claim 1 , wherein the nucleotide sequence comprises the nucleotide sequence set forth in SEQ ID NO: 15.
12. The method of claim 2 , wherein NBB1 is overexpressed.
13. The method of claim 2 , wherein A622 is overexpressed.
14. The method of claim 2 , wherein QPT is overexpressed.
15. The method of claim 2 , wherein PMT is overexpressed.
16. The method of claim 2 , wherein MPO is overexpressed.
17. The transgenic Nicotiana plant of claim 6 , wherein the plant is a Nicotiana tabacum plant.