Method for increasing a nicotinic alkaloid in a
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. The present invention is directed to a method for increasing a nicotinic alkaloid in a nicotiana plant by introducing a mutation into the gene encoding the NbTF7 transcription factor. The present invention is also directed to mutagenized plants produced by this method and products made from these plants.
1. A method for increasing a nicotinic alkaloid in a Nicotiana plant, comprising:
(a) genetically engineering a population of plant cells to eliminate production of a protein encoded by the nucleotide sequence set forth in SEQ ID NO: 14, comprising introducing one or more mutations into the region of SEQ ID NO: 14 that encodes a polypeptide having the amino acid sequence set forth in SEQ ID NO: 16; and
(b) detecting and selecting a target mutated plant cell or a plant derived from such a cell, wherein the target mutated plant cell or plant has a mutation in a gene encoding a transcription factor that negatively regulates nicotinic alkaloid biosynthesis and wherein the targeted plant cell or plant has an increased nicotinic alkaloid content as compared to a control plant cell or plant.
2. The method of claim 1 , wherein the genetic engineering further comprises introducing into the population of cells a reagent comprising a recombinagenic oligonucleobase.
3. The method of claim 1 , wherein the genetic engineering further comprises introducing into the population of cells a reagent comprising a targeted nuclease.
4. A mutated Nicotiana plant having reduced expression of a transcription factor that negatively regulates nicotinic alkaloid biosynthesis and increased nicotinic alkaloid content, as compared to a control plant, produced by the method of claim 1 .
5. The mutated plant of claim 4 , wherein the plant is a Nicotiana tabacum plant.
6. A product having an increased level of a nicotinic alkaloid comprising the mutated plant of claim 4 , or portions thereof, as compared to a product produced from a control plant.
7. Seeds from the mutated plant of claim 4 , wherein said seeds comprise said mutation.
8. The product of claim 6 , 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. The method of claim 1 , wherein the genetic engineering further comprises overexpressing within the Nicotiana plant at least one of NBB1, A622, quinolate phospho-ribosyltransferase (QPT), putrescine-N-methyltransferase (PMT), and N-methylputrescine oxidase (MPO).
10. The method of claim 1 , wherein the genetic engineering further comprises overexpressing within the Nicotiana plant at least one transcription factor that positively regulates nicotinic alkaloid biosynthesis.
11. The method of claim 1 , wherein the genetic engineering further comprises suppressing the expression of at least one additional transcription factor that negatively regulates nicotinic alkaloid biosynthesis.
12. The product of claim 6 , wherein the product is a cigarette.
13. The product of claim 6 , wherein the product is cigarette tobacco.
14. The product of claim 6 , wherein the product is cigar tobacco or a cigar.
15. The product of claim 6 , wherein the product is, pipe tobacco, chewing tobacco, or snuff.