Designing, construction and transformation of silencing constructs for F3´H and F3´5´H genes in grapevine cv. Shiraz

Document Type : Full Paper


1 Post Graduate Student and Professor, University College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran

2 . Post Graduate Student and Professor, University College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran

3 . Associate Professor, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran

4 . Researcher and Professor in CSIRO Plant Industries, Adelaide, Australia


 In order to study F3´5´H and F3´H function and their intraction in the grapevine flavonoids biosynthetic pathway, designing and constraction of suitable constructs encoding ihpRNA was performed and transgenic lines containing silencing construct of F3´H and simultaneous silencing construct of F3´5´H and F3´H were created. For this purpose, fragments were selected from both genes and inserted into pHANNIBAL vector as inverted repeats. Using binary vector system, each construct inserted into p27 mod GFP 4a vector and were used for Agrobacterium transformation. Embryogenic calli, drived from anther culture, were co-cultivated with Agrobacterium for transformation. Expression of GFP gene was detected in transgenic embryogenic calli, 5 days after inoculation using microscope. Finally, 42 and 34 lines were generated from single (F3´H) and simultaneous silencing (F3´5´H and F3´H) cultures, respectively. PCR analysis confirmed 36 single (F3´H) silencing line and 27 simultaneous silencing (F3´5´H and F3´H) lines, showing high efficacy of regeneration and transformation methods.



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