The effects of matrix attachment regions on RNA-mediated virus resistance

RNA-mediated virus resistance in plants works through the post-transcriptional gene silencing (PTGS) of a viral transgene. This method produces high levels of virus resistance, but resistance can be unstable in later generations. It was hypothesized that flanking a construct for virus resistance with DNA elements called matrix attachment regions (MARs) would increase the frequency of virus resistant lines and provide greater stability of resistance in later generations.; Two populations of transgenic tobacco were developed. The MAR population was transformed with the tomato spotted wilt virus (TSWV) nucleocapsid (N) gene flanked with MARs, and the nonMAR population was transformed with a similar construct without MARs. The MAR population had a significantly higher frequency of resistance in the R1 generation, and resistance was significantly more stable over four generations. Both MAR and nonMAR lines with resistance in growth chamber tests had excellent resistance to TSWV in field trials.; It was hypothesized that loss of resistance was caused by transcriptional silencing (TGS) of the transgene. The resistant R1 and the susceptible R4 generations of two lines that lost resistance were examined to determine if TGS had occurred. Nuclear run-on assays showed that transgene transcription was reduced or suppressed, indicating that loss of resistance was caused by TGS. Since loss of resistance occurs less frequently in the MAR population, MARs may be acting to prevent TGS. By providing a high frequency of stable virus resistance, the use of MARs could reduce the cost of developing virus resistant transgenic plants.; Some viruses are able to suppress PTGS and could potentially interfere with transgenic resistance. Infection with some isolates of potato virus Y, tobacco etch virus, and cucumber mosaic virus could suppress PTGS in a TSWV-resistant line, but there was significant variation in the ability of different isolates to suppress PTGS. Isolates of potato virus Y and tobacco etch virus were able to cause TSWV-resistant plants to become susceptible to TSWV. This may have implications for the efficacy of this resistance in the field.