<p style="text-align: justify;"><b>The Introduced DNA</b></p>

<p style="text-align: justify;">Since PWC16 is a product of mutagenesis and conventional seed increase techniques there was no introduction or incorporation of heterologous DNA into the plant genome. The tolerance to imazethapyr is due to a single point mutation in the AHAS encoding gene such that the amino acid sequence of the mutated enzyme differs by one amino acid from that of the unmodified enzyme. This single change in the amino acid sequence alters the binding site on AHAS to imazethapyr, such that the herbicide cannot inhibit the enzyme&rsquo;s activity. This mutation of the AHAS gene was identified by sequencing the gene, which showed the single nucleotide change in the coding sequence for AHAS.</p>

<p style="text-align: justify;"><b>Genetic Stability of the Trait</b></p>

<p style="text-align: justify;">Segregation analysis of crosses with PWC16 demonstrated that a single semi-dominant gene was inherited. All F1 progeny are resistant to imazethapyr, while F2 progeny segregate in a 1:2:1 ratio (herbicide resistant: intermediate resistant: susceptible to herbicide).</p>

<p style="text-align: justify;"><b>Expressed Material</b></p>

<p style="text-align: justify;">The modified AHAS gene, conferring tolerance to imazethapyr, is under control of the native AHAS promoter and is believed to be constitutively expressed. Whole plant tolerance to this herbicide was expressed in PWC16 rice.</p>

<p style="text-align: justify;">In the unmodified plant, the levels of valine, leucine and isoleucine are regulated by feedback inhibition. A mutation in the AHAS enzyme could affect the regulation of the biosynthesis of these amino acids. Data were submitted to show that the activity of the modified AHAS was not altered and did not affect feedback inhibition and therefore, the regulation and levels of these amino acids. The amino acid composition of PWC16 was compared to that of commercial cultivars and found to be equivalent in terms of the levels of valine, leucine and isoleucine.</p>
