The Development of an Efficient Multipurpose Bean Pod Mottle Virus Viral Vector Set for Foreign Gene Expression and RNA Silencing

摘要

Plant viral vectors are valuable tools for heterologous gene expression, and because of virus-induced gene silencing (VIGS), they also have important applications as reverse genetics tools for gene function studies. Viral vectors are especially useful for plants such as soybean (Glycine max) that are recalcitrant to transformation. Previously, two generations of bean pod mottle virus (BPMV; genus Comovirus) vectors have been developed for overexpressing and silencing genes in soybean. However, the design of the previous vectors imposes constraints that limit their utility. For example, VIGS target sequences must be expressed as fusion proteins in the same reading frame as the viral polyprotein. This requirement limits the design of VIGS target sequences to open reading frames. Furthermore, expression of multiple genes or simultaneous silencing of one gene and expression of another was not possible. To overcome these and other issues, a new BPMV-based vector system was developed to facilitate a variety of applications for gene function studies in soybean as well as in common bean (Phaseolus vulgaris). These vectors are designed for simultaneous expression of multiple foreign genes, insertion of noncoding/antisense sequences, and simultaneous expression and silencing. The simultaneous expression of green fluorescent protein and silencing of phytoene desaturase shows that marker gene-assisted silencing is feasible. These results demonstrate the utility of this BPMV vector set for a wide range of applications in soybean and common bean, and they have implications for improvement of other plant virus-based vector systems.