Cloning and functional study of the alkaline/neutral invertase gene PtoNIN1 in Populus tomentosa

  Objective  Sucrose invertase, as a key enzyme in plant sucrose metabolism, plays an important role in plant growth and development. In this study, PtoNIN1, an alkaline/neutral invertase, was selected to do detailed analysis, including gene cloning, bioinformatics, gene expression and genetic transformation analysis, and these results were helpful for clarifying the mechanism of sucrose metabolism in Populus tomentosa.

  Method  PtoNIN1 was obtained through the homologous cloning of PtrNIN1 in P. trichocarpa, and then bioinformatics analysis for PtoNIN1 was carried out. Gene expression of PtoNIN1 was conducted at different organs (leaf, stem, root and mature leaves) and different developmental stages of male and female flower buds. And overexpression vector of PtoNIN1 was constructed to carry out genetic transformation research on the model plant Arabidopsis thaliana.

  Result  The coding region of PtoNIN1 was 2 073 bp in length. The PtoNIN1 protein contained 690 amino acids, and its molecular mass was 77.80 kDa. PtoNIN1 contained a specific domain of glycosyl hydrolase supergene family 100. The phylogenetic analysis showed that it is the member of mitochondrial-type invertase cluster. PtoNIN1 was widely expressed in the plant, including roots, stems, leaves, mature leaves, male and female flower buds. Thereinto, the development stage of male and female flower buds had an influence on the expression of PtoNIN1. Genetic transformation studies in Arabidopsis showed that overexpression of PtoNIN1 significantly increased the fresh mass of rosette leaves, stems and siliques. The sucrose contents in stems and siliques of transgenic plants were also significantly increased. And higher expression patterns of other sucrose metabolism-related genes were found in transgenic plants when compared with wild-type plant.

  Conclusion  PtoNIN1 is the member of mitochondrial-type invertases in P. tomentosa. It is widely expressed in various tissues of poplar, and has a specific expression in the early developmental stage of male and female inflorescences. Overexpression of PtoNIN1 in Arabidopsis can significantly increase plant biomass accumulation. This study provides a reference for the functional verification of other invertases in poplar, and lays a foundation for revealing the sucrose metabolic process of poplar.