Figure 4.

Soybean transgenic plants expressed the mRNA and protein from the Forrest allele of GmRLK18-1 , the RLK at the Rfs2/Rhg1 locus. Panel A shows PCR from leaf samples of progeny plants derived from a primary transgenic event 6B3-7D2(1) with TMD1 primers. Lanes contain transgenic plants 1 to 13. The arrow shows the double band for Gm18RLK-1-a positive sample at 314 bp for lines 1,3–5, 7,8,10 and 12. M was the marker; H was the no DNA (water) control; P was the Rhg1 plasmid pSBHB94; X5 was the control plant. Panel B shows PCR from cDNA leaf samples of sixteen transgenic lines derived from event 6B3-7D2(1) with HRM primers. Green lines are from transgenic plants. Red melt curve was a resistant control blue line was a susceptible control. Panel C shows a Western of a 2D gel from roots of a transgenic plant probed with the anti-RLK peptide antibody. An alloprotein at pI 8.42 and 92.41 kDa was found in the non transgenic cv X5 but the presence of the Forrest alloprotein at pI 8.44 and and 92.39 kDa was found in transgenic plants derived from event 6B3-7D2(1) expressing GmRLK18-1-a. GmRLK18-1 was shown to be a very low abundance protein impossible to visualize without immune-staining.