| RNA silencing is identified as a general and ancient defense mechanism of plants against virus infections. As a genome defense mechanism, RNA silencing has been termed as post transcriptional gene silencing (PTGS) in plants. Recent studies have revealed that this mechanism is conserved among various eukaryotic organisms to counter the proliferation of alien sequences, such as transposable elements, transgenes and viruses, by specifically degrading mRNAs homologous to the target genes. RNA-mediated virus resistance is induced by transgenes derived from genes of virus and regarded as a potential strategy with application value in plant resistance breeding against viruses, which has the advantages of extreme resistance (immune), high biosafety and long duration. VIGS is a natural plant antiviral mechanism.In this study,we take the transgenic tobacco K58 which has the IR from PVY~N CP to research the influence of IR-mediatedly transitive distance toward upstream of PVY~N CP, and detect the function of RDR1 in this process by the VIGS vector. This is beneficial to produce proof of nurturing multi antiviral transgenic plants and maintaining long-playing RNA-mediated disease resistance. The main results are as follows:1.We clone the 373bp Nt-RDR1 fragment from Nicotiana tabacum NC89,which is conservative with N. benthamian(97.32%). The fragments were inserted into viral vector TRV and PVX, we name the vectors as TRV-RP and PVX-RP which are used to silence the RDR1.2. These vectos were transformed into the Agrobacterium strain GV3101, N. benthamian and K58 were inoculated by infiltration with culture of Agrobacterium GV3101. Plants were grown in glass house at 20°C and 14-h-light/10-h-dark cycles. After 20 days RNA was extracted from plants and cDNA was composition by RT-PCR. After the detection by Real Time PCR and Northern Blotting, we found that expression of the RDR1 gene was obviously increased in the plants which were treated by TRV vector, and obviously reduced in the plants which were infected by TRV-RP and PVX-RP vector. But in the new leaves of K58 we could not detect the TRV-RP and PVX-RP vector, and the RDR1 gene was not silenced. However we can detect it in the N. benthamian.3.We take PVY~N infect the K58 which dealt with TRV-RP and TRV vector at 15dpi. Plants were grown in glass house at 20°C and 14-h-light/10-h-dark cycles. After 4dpi, RNA was extracted from plants and cDNA was composition by RT-PCR. After the detection by Real Time PCR, plants with lower accumulation of RDR1 transcripts(infected by TRV-RP) showed much higher accumulation levels of PVY than the plants treated by TRV vector. When we detected the influence of transitive distance toward upstream by the Real Time PCR primers A,B,C,D,HC and EF1α,the result was that the infected leaves which treated by TRV-RP had a lower level of silencing with every primer. The level of new leaves was similar with the control plants.4. We take the K58 infected by PVY~N in glass house at 15°C, 20°C, 25°C, 30°C, 35°C and 14-h-light/10-h-dark cycles. After 4dpi, RNA was extracted from plants and cDNA was composition by RT-PCR. When we detected the influence of distance of transitive silencing toward upstream by the Real Time PCR primers A,B,C,D,HC and EF1α, the result shown that the level of silenced transitively distance was 30℃> 25℃> 20℃> 15℃> 35℃.
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