Functional Analysis Of PtrPRP From Poncirus Trifoliata In Cold Tolerance

Poncirus trifoliata (L.)Raf is the most widely used root stock in citrus planting, This study is based on a low temperature responsive genes PtrPRP, which is previously cloned using SSH (Suppression subtractive hybridization), but the functional of cold stress is not clear. For further characterization of the function of PtrPRP, its expression patterns under various stresses (low temperature, dehydration, salinity, and exogenous ABA) were analyzed. In addition, PtrPRP-RNAi was introduced into the trifoliate orange via Agrobacterium-mediated transformation. The main results are as follows:1. Real-Time PCR analysis suggested that PtrPRP was induced by multi stimuli (low temperature, dehydration, salinity and exogenous ABA treatments), and strongly induced by low temperature. Besides that, Tissue-specific expression profiles showed that PtrPRP most accumulated in leaf, and lowest in root.Subcellular localization analysis revealed that it may localize in the plasma membrane.2.31positive transgenic poncirus plants were identified by PCR, which transformed with PtrPRP-RNAi. Then RT-PCR detected that the expression of PtrPRP in the selected transgenic lines (51#and52#) was lower compare to wild type. However, In the0℃treatment for the WT and51#,52#. The measurement of Electrolyte leakage and MDA determined that51#and52#were higher than WT. in addition, DAB and NBT staining demonstrated that both transgenic lines were markedly stronger than WT, suggesting that both transgenic plants accumulated more H2O2and O2. Consistently the expression and the content of proline in both transgenic lines were also lower than WT.Taken together, we can determine that PtrPRP gene have the function of responding to cold stress.