Differential Proteome Analysis Of Hybrid Rice In Response To Salt Stress

Two hybrid rice combinations, shanyou10 (salt-tolerant) and liangyoupeijiu (salt-sensitive), were used for this experiment. Two-dimentional polyacrylamide gel electrophoresis and MALDI-TOF-MS/MS techniques were empolyed to isolate and characterize salt stress responsive proteins of hybrid rice seedlings in early period of 10-day old or at three-leaf-stage (18-day old) under normal and salt stress (100mM NaCl) conditions The results showed that1. Seeds of two hybrid rice combinations were planted in germination box containing two-layer paper moisted with water (control) or 100 mM NaCl solution, respectively. After treatment for ten days, shoots and roots were collected, and the total proteins are extracted using TCA acetone precipitation. The total proteins were separated using 2-DE with linear pH 4-7 IPG strip. It is indicated by PDQuest software analysis that the matching percent of four 2-D pattern reached more than 90%. Twenty-five or forty up-regulated spots were significantly found in shanyou10 or in liangyoupeijiu leaves compared with the control. Six of them were selected and identified by MALDI-TOF-MS/MS, Only the two uniproteins, ribulose-1,5-bisphosphate carboxylase/oxygenase and enolase, were identified. Intriguingly, ninety-six or sixty-eight up-regulated spots and seventy-three or eighty-five down-regulated spots were observed in shanyou10 or in liangyoupeijiu roots under salt stress, respectively . Eleven of them were selected and identified by MALDI-TOF-MS/MS. Eight uniproteins, including pathogenesis-related thaumatin-like protein, putative antifungal zeamatin-like protein, GH18_narbonin, Ascorbate peroxidase, Glyoxalase, GH18_hevamine_XipI_class_III, Glycosyl hydrolases, Glutathione S-transferase, were obtained. These proteins are involved in carbohydrate and energy metabolism regulation, disease control, ROS detoxification and protein synthesis and localization.2. Eighteen-day old rice seedlings were treated with 100mM NaCl for a week under solution culture condition. shoots and roots were collected, and the total proteins are extracted using TCA acetone precipitation. The total proteins were separated using 2-DE with linear pH3-10 or pH5-8 IPG strip.condition, respectively. Based on the results of three repeats, 2-D pattern of two hybrid rice combinations was shown better reproducibility. PDQuest software analysis showed that the matching percent of four 2-D pattern reached more than 90%. In 2-D pattern with pH3-10, there were six protein spots with up-regulated two fold in shanyou10, and fifteen spots in liangyoupeijiu under the salt stress compared with the control. Nine of them were selected and identified by MALDI-TOF-MS/MS. three uniproteins were identified, Including Ribulose-1,5-bisphosphate carboxylase/oxygenase triosephosphate isomerase (TIM), chaperonin GroEL. In the 2-DE pattern with pH5-8, there are thirty-seven protein spots in shanyou10 and thirty-six in liangyoupeijiu with up-regulated expression under salt stress, Twenty-four of them appeared to be differentially expressed. Six of them were selected and identified by MALDI-TOF-MS/MS. Three uniproteins were obtained, inculding Ribulose bisphosphate carboxylase, Ribulose bisphosphate carboxylase/oxygenase Activase and Ribosome recycling factor. These proteins are mainly involved in photosynthesis, energy metabolism and protein synthesis regulation.