Simple gene silencing using the trans‐acting si RNA pathway

Summary In plants, particular micro‐ RNA s (mi RNA s) induce the production of a class of small interfering RNA s (si RNA ) called trans‐acting si RNA (ta‐si RNA ) that lead to gene silencing. A single mi RNA target is sufficient for the production of ta‐si RNA s, which target can be incorporated into a vector to induce the production of si RNA s, and ultimately gene silencing. The term mi RNA ‐induced gene silencing ( MIGS ) has been used to describe such vector systems in Arabidopsis . Several ta‐si RNA loci have been identified in soybean, but, prior to this work, few of the inducing mi RNA s have been experimentally validated, much less used to silence genes. Nine ta‐si RNA loci and their respective mi RNA targets were identified, and the abundance of the inducing mi RNA s varies dramatically in different tissues. The mi RNA targets were experimentally verified by silencing a transgenic GFP gene and two endogenous genes in hairy roots and transgenic plants. Small RNA s were produced in patterns consistent with the utilization of the ta‐si RNA pathway. A side‐by‐side experiment demonstrated that MIGS is as effective at inducing gene silencing as traditional hairpin vectors in soybean hairy roots. Soybean plants transformed with MIGS vectors produced si RNA s and silencing was observed in the T1 generation. These results complement previous reports in Arabidopsis by demonstrating that MIGS is an efficient way to produce si RNA s and induce gene silencing in other species, as shown with soybean. The mi RNA targets identified here are simple to incorporate into silencing vectors and offer an effective and efficient alternative to other gene silencing strategies.