Compositions and methods based on pmt engineering for producing tobacco plants and products having altered alkaloid levels

A tobacco plant, or part thereof, comprising:

• (a) a first knockout mutant allele in a putrescine N-methyltransferase (PMT)1a gene (PMT1a), wherein a wildtype allele of the PMT1a gene encodes the polypeptide of SEQ ID NO:
• 12, and wherein the first knockout mutant allele in the PMT1a gene comprises the sequence of SEQ ID NO:
• 432;
• and
• (b) a second knockout mutant allele in a PMT1b gene, wherein a wildtype allele of the PMT1b gene encodes the polypeptide of SEQ ID NO:
• 11;
• and
• (c) a third knockout mutant allele in a PMT2 gene, wherein a wildtype allele of the PMT2 gene encodes the polypeptide of SEQ ID NO:
• 13;
• and
• (d) a fourth knockout mutant allele in a PMT3 gene, wherein a wildtype allele of the PMT3 gene encodes the polypeptide of SEQ ID NO:
• 14;
• and
• (e) a fifth knockout mutant allele in a PMT4 gene, wherein a wildtype allele of the PMT4 gene encodes the polypeptide of SEQ ID NO:
• 15,
• wherein the cured tobacco plant material comprises a tobacco leaf comprising a nicotine level less than 0.25% of the nicotine level from a control tobacco plant lacking the first knockout mutant allele, the second knockout mutant allele, the third knockout mutant allele, the fourth knockout mutant allele, and the fifth knockout mutant allele when grown and processed under comparable conditions.