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

<p style="text-align: justify;">The synthetic <i>pat</i> gene was modified to provide codons preferred by plants without changing the amino acid sequence of the PAT enzyme. The PAT protein under study was found to have a similar molecular weight to the bacterial PAT protein.</p>

<p style="text-align: justify;">Southern blot analyses indicated that the DNA was integrated in the original transformant at one insertion site and the insertion contained 2 linked copies of the <i>pat</i> gene. Additional Southern blot analyses were performed to demonstrate that plasmid backbone sequences outside of the T-DNA region were not incorporated into the host genome.</p>

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

<p style="text-align: justify;">HCN92 was several generations removed from the original transformant and comparisons between the original transgenic plant (Topas19/2) and HCN92 showed no differences in the presence and expression of the <i>pat</i> and <i>neo</i> genes, nor in the insertion site.</p>

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

<p style="text-align: justify;">Constitutive expression of the PAT protein was detected in roots, leaves, buds and seeds. Seed expression levels ranged from 150 to 223 ng/g of seed tissue. PAT protein was not detected in stem tissue, protein extracts from the pollen, or unprocessed honey. Maximum expression was 0.001% of total plant protein.</p>

<p style="text-align: justify;">The <i>neo</i> gene was linked to a weak constitutive promoter, and expression was consistently stronger in root tissue, but was also observed in buds, leaves, and crude seed samples. The enzyme was not detected in unprocessed honey or pollen samples and was inactivated during processing of canola seed into feed ingredients.</p>
