Screening and Validation of Rice OsAAP6 Interaction Protein

Abstract

The protein content of rice seeds is an extremely important quality trait, but its genetic basis and molecular regulatory mechanism are still unclear. This study focuses on a positive regulatory gene OsAAP6 of grain protein content in rice. Proteins that interact with OsAAP6 were screened using yeast two hybrid experiments, and validated using in vivo point-to-point experiments and bimolecular fluorescence complementarity tests (BiFC). The main results are as follows: 98 positive colonies that may interact with OsAAP6 were screened from a rice cDNA library using yeast two hybrid technology. After sequencing and analysis, 40 proteins that may interact with OsAAP6 were ultimately obtained. Through comparative analysis, three proteins (Pyruvate phosphate dikinase (PPDR), WRKY, and GreA) were selected from these 40 proteins that may interact with each other. In vivo point-to-point experiments in yeast and bimolecular fluorescence complementarity (BiFC) experiments in rice were used to further verify that PPDK, WRKY, and GreA can interact with OsAAP6 protein, respectively. Therefore, the results of this study will provide important clues for further revealing the molecular mechanism by which the OsAAP6 gene regulates grain protein content.