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Expression of the ZmDEF1 gene and α-amylase inhibitory activity of recombinant defensin against maize weevils

Plant defensins are multifunctional small cysteine-rich proteins. They are active against fungi, bacteria, and many viruses, and they inhibit trypsin and α-amylase activities. In this study, we expressed the maize defensin gene (ZmDEF1) in tobacco seeds in order to establish the basis for generating transgenic maize plants resistant to weevils. | Turkish Journal of Biology Turk J Biol (2017) 41: 98-104 © TÜBİTAK doi:10.3906/biy-1512-64 http://journals.tubitak.gov.tr/biology/ Research Article Expression of the ZmDEF1 gene and α-amylase inhibitory activity of recombinant defensin against maize weevils 1 2 3 2 4, Thi Xuan Thuy VI , Hoang Duc LE , Vu Thanh Thanh NGUYEN , Van Son LE , Hoang Mau CHU * 1 Tay Bac University, Son La, Vietnam 2 Institute of Biotechnology, Vietnam Academy of Science and Technology, Hanoi, Vietnam 3 Thai Nguyen University of Sciences, Thai Nguyen, Vietnam 4 Department of Genetics and Modern Biology, Thai Nguyen University of Education, Thai Nguyen, Vietnam Received: 24.12.2015 Accepted/Published Online: 21.07.2016 Final Version: 20.02.2017 Abstract: Plant defensins are multifunctional small cysteine-rich proteins. They are active against fungi, bacteria, and many viruses, and they inhibit trypsin and α-amylase activities. In this study, we expressed the maize defensin gene (ZmDEF1) in tobacco seeds in order to establish the basis for generating transgenic maize plants resistant to weevils. The ZmDEF1 gene was isolated from Maison, a Vietnamese local maize cultivar, which is well known for having the highest resistance to weevils among other local cultivars. The ZmDEF1 gene was cloned into a binary vector, pBetaPhaso-dest, which carries phaseolin, a seed-specific promoter, to direct defensin expression in tobacco seeds. We obtained 13 transgenic tobacco lines from Agrobacterium-mediated transformation and regeneration, called the T0 generation. T0’s seeds (called the T1 generation) were collected and analyzed for ZmDEF1 gene expression. Reverse-transcription polymerase chain reaction (RT-PCR) results showed that 4 out of 13 lines (T1-1, T1-3, T1-10, and T1-17) expressed the ZmDEF1 gene at the transcriptional level. These lines were further analyzed by real-time RT-PCR until their transcript expression could be identified. The results showed that the line T1-17 expressed it .