Construction and Identification of TRV-mediated VIGS Transformation System of Iris sanguinea
To analyze gene function in plants lacking stable genetic transformation system,virus-induced gene silencing(VIGS)was needed,and Iris sanguinea,a monocotyledon,was selected as materials.The specific fragment of IsPDS gene was isolated and the VIGS recombinant vector pTRV2-IsPDS was constructed and leaves were infected by injection.The results showed that the most effective infection was achieved by injecting its leaf veins with syringes when the OD600 values of the resuspension were adjusted to 0.8-1.0 after the those of pTRV1 and pTRV2-IsPDS were adjusted to 1.8-2.0.The experiment was conducted when outdoors temperature was 15-20℃from 6 p.m.to 8 p.m.,and a 1 mL syringe needle pricking the outer epidermis of I.sanguinea leaves and 1 mL of heavy suspension slowly injected along parallel veins into its vascular bundles.A clear albino phenotype might appear after about 14 days.TRV1 and TRV2 virus vectors were detected in the plants with phenotypic changes and in the no-load group.The expression of IsPDS in the albino plants was significantly lower than that in the no-load group and the control group.With the concentration of agrobacterium tumefaciens carrying virus vector increased in the preparation of infection solution,the infection efficiency of the whole experiment was improved,and no shading was needed after inoculation.
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维普
