COMPARISON STUDY OF RESISTANCES AGAINST ARTIFICIAL INOCULATION OF SCMV IN TRANSGENIC SUGARCANE DEVELOPED BY PDR AND RNAi METHODS
Weny Nailul Hidayati1, Suvia Widyaningrum1, Retnosari Apriasti2, Bambang Sugiharto2,3*
1 Magister Biotechnology, University of Jember, Jln. Kalimantan 37, Jember 68121, Indonesia
2Laboratory of Molecular Biology and Biotechnology, Center for Development of Advanced Sciences and Technology (CDAST), University of Jember, Jln. Kalimantan 37, Jember 68121, Indonesia
3Department of Biology, Faculty of Mathematics and Natural Sciences, Jember University,
Jln. Kalimantan 37, Jember 68121, Indonesia
Abstract
Background: SCMV (Sugarcane Mosaic Virus) is a disease that cause reduction in growth and productivity of sugarcane. Infection of SCMV show yellowish spots symptomatic sugarcane leaves that indicates a decrease in the concentration of chlorophyll. Infection by the virus causes chloroplast damage, decreased photosynthesis and a sugar productivity loss of approximately 20–50% [1, 2]. Aims : to compare resistences of transgenic sugarcane derived from PDR and RNAi method against artificial infection of SCMV Materials and Methods: the experiment was conducted by cultivation of PDR and RNAi-transgenic sugarcane in green house for 3 weeks. Artificial inoculation of SCMV was performed on the leaves of 3 weeks transgenic sugarcane. The resistences of transgenic sugarcane against SCMV infection were observed by the presence of yellowish spot and by analysis of RT-PCR and western blot. Results: One of the efforts to overcome the SCMV infection is the development of resistance against SCMV through genetic transformation. The transgenic sugarcane resistant to SCMV was successfully developed by Pathogen Derived Resistence (PDR) and RNA interference (RNAi) approaches using coat protein mediated resistances. Both of these techniques have the ability to suppress virus infection, but there is no study on their effectiveness. In this study, the transgenic sugarcane were grown and treated with SCMV inoculation test to determine and compare the resistances. The symptomatic observation and molecular analysis such as PCR, Western blot and RT-PCR were conducted to determine the effectiveness of the transgenic sugarcane. The PCR analysis confirmed that the transgenic sugarcane were genetically stable inserted by the targeted coat protein DNA at ± 700bp. The level of expression of coat protein genes was analyzed using Western blot and showed the results of protein size ± 30 Kda. Conclusions:Artificial inoculation using SCMV virus sap fluid showed that non-transgenic sugarcane have yellowish spots symptoms on the leaves, while transgenic sugarcane did not show the symptoms. This results showed that transgenic sugarcane developed by PDR and RNAi methods inhibit the replication of the SCMV virus when it is infected. References 1. Putra LK, Kristini A, Achadian EM, Damayanti TA (2014) Sugarcane streak mosaic virus in Indonesia: distribution, characterisation, yield losses and management approaches. Sugar Tech 16:392–399. https ://doi.org/10.1007/s1235 5-013-0279-9 2. Akbar S, Tahir M, Wang M-B, Liu Q (2017) Expression analysis of hairpin RNA carrying sugarcane mosaic virus (SCMV) derived sequences and transgenic resistance development in a model rice plant. BioMed Res Int. https://doi.org/10.1155/2017/16461 40
Keywords: PDR, RNAi, Sugarcane Mosaic Virus, resistances, artificial inoculation.
Topic: Environmentally Sustainable Agriculture