Composite plants: A novel method for gene screening in cowpea

Background/Question/Methods

Legumes such as cowpea (or black-eyed pea) are a notable source of protein for both humans and livestock and are of significant interest to global nutrition. Cowpea thrives in sandy soils and arid climates, such as those in sub-Saharan Africa where drought due to climate change is expected to worsen. Striga, a parasitic plant genus that attacks healthy cowpea roots, threatens cowpea’s growth in Africa, with up to 70% crop yield loss. Fortunately, some cowpea strains are known to have developed resistance to Striga. The purpose of this study was to develop a high-efficiency method for testing Striga resistance genes in cowpea. Ex vitro composite plant technology allows gene overexpression and regeneration of transgenic roots, eliminating the need for a wholly transgenic organism, thus maximizing screening efficiency. In this study, cowpea was transformed using the composite plant method with the plant pathogen Agrobacterium rhizogenes as a gene transfer vector. A. rhizogenes was selected to contain pCAMBIA1300, a plasmid containing GFP to indicate successful transformation. Composite plant setups were performed at 21, 23, and 25°C to determine optimal temperature for highest transgenic root yield. A. rhizogenes density and hypocotyl wounding length were also tested. Individuals were inspected for GFP after successful root growth in the composite system. 

Results/Conclusions

The 21°C treatment showed to be significantly more efficient (approaching 25% of roots transgenic) when compared to the 25°C treatment (7%) and the 23°C treatment (10%). A. rhizogenes OD (optical density) was not found to affect transformation efficiency.  These results show that successful generation of transgenic roots for testing Striga interaction will be much more efficient at 21°C. This is believed to be a function of plant stress, as successful infection by A. rhizogenes requires increased susceptibility of cowpea to bacterial infection. Additionally, the gene construct is thought to be more stable at lower temperatures. As a result of temperature optimization, candidate resistance genes may be tested against Striga using a greater number of host roots, where transgenic roots will be resistant to the Striga parasite. Successfully transformed plants will be of significant interest to agriculture where Striga is rampant. Knowledge of the resistance gene will allow hardier, more nutritious strains of cowpea to grow in drought-prone areas. Additionally, this study shows that composite plant technology is appropriate for studying gene function in cowpea roots.