Start Date: Jan 01, 2013
End Date: Dec 31, 2013
We have over 100 independent Coker 312 callus lines transformed with an intron-containing hair-pin (ihp) construct of desoxyhemigossypol-6-O-methyltransferase (dHG-6-OMT) sequence from Coker 312 driven by the CaMV35S promoter to reduce the expression of dHG-6-OMT throughout the plant. We will generate transgenic cotton (G. hirsutum) plants from these transformed lines. We will confirm that the transgenic plants contain the RNAi construct by Southern hybridization and by quantitative Reverse Transcriptase PCR (q-RT-PCR) of dHG-6-OMT mRNA. We will determine if the dHG-6-OMT mRNA is significantly reduced as expected in root tip tissues of young seedlings of G. hirsutum and we expect that the dHG-6-OMT mRNA levels will to be even lower in mature glanded tissues. In addition, we will analyze root tissues for the presence of methylated phytoalexins; again, we expect a decrease of methylation to correlate with the RNAi suppression of dHG-6-OMT transcripts.