Ectopic expression of antifreeze protein gene from Ammopiptanthus nanus confers chilling tolerance in maize
Yuanyuan Zhang,
Yang Cao,
Hongying Zheng,
Wenqi Feng,
Jingtao Qu,
Fengling Fu,
Wanchen Li,
Haoqiang Yu
Affiliations
Yuanyuan Zhang
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China; College of Life Science & Biotechnology, Mianyang Teachers’ College, Mianyang 621000, Sichuan, China
Yang Cao
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China
Hongying Zheng
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China
Wenqi Feng
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China
Jingtao Qu
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China
Fengling Fu
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China
Wanchen Li
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China; Corresponding authors.
Haoqiang Yu
Key Laboratory of Biology and Genetic Improvement of Maize in Southwest Region, Ministry of Agriculture, Maize Research Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan, China; Corresponding authors.
Improved chilling tolerance is important for maize production. Previous efforts in transgenics and marker-assisted breeding have not achieved practical results. In this study, the antifreeze protein (AnAFP) from the super-xerophyte Ammopiptanthus nanus was aligned to KnS-type dehydrins. Phosphorylation in vitro and subcellular localization showed that AnAFP was phosphorylated by maize casein kinase II and translocated from nucleus to cytoplasm under chilling stress. AnAFP also increased lactate dehydrogenase activity. A parent line of commercial maize hybrids was transformed with the AnAFP gene. Based on thermal asymmetric interlaced PCR, one hemizygous and two homozygous integration sites were identified in one T1 line. Ectopic expression of AnAFP in transgenic lines was confirmed by quantitative real-time PCR, RNA-seq, and Western blotting. After chilling treatment, the transgenic lines showed significantly improved phenotype, higher kernel production, survival rate and biomass, and lower relative electrolyte leakage and superoxide dismutation than the untransformed line. Thus, ectopic expression of AnAFP gene improved chilling tolerance in the transgenic lines, which could be used to apply for further safety assessment for commercial breeding.
