Function Characterization Of Poncirus Trifoliata (L.) Raf. In Cold Resistance

The key to citrus genetic engineering is the identification of stress resistance genes. The gene, PtrbHLH, was induced by low temperature. The cold tolerance were improved when the PtrbHLH gene was overexpressed in tobacco and lemon. In this study, PtrbHLH-RNAi and PtrbHLH were introduced into the trifoliate orange(Poncirus trifoliata L. Raf) and pomelo(Citrus grandis(Linn.) Osbeck) via Agrobacterium-mediated transformation, in order to further identify of the function of PtrbHLHin cold resistance. The main results are as follows:1.Thirty positive transgenic plants were identified by PCR in poncirus transformed with PtrbHLH-RNAi. RT-PCR analysis showed that the expression level of PtrbHLH in the selected transgenic lines(#5and#7) were lower than in WT. Enzyme assay results showed that POD and CAT enzyme activity in two transgenic lines (#5and#7) was significantly lower than WT. With0℃low temperature treatment,#5and#7exhibited worse wilting than WT. Conductivity measurement results showed that#5and#7were significantly higher than WT. DAB staining results showed that the staining of the transgenic lines#5and#7was stronger than WT, indicating that the transgenic plants accumulated more H2O2. Enzyme assay results showed that treatment at low temperature. POD activity in the#5and#7was lower than WT.2.Four positive transgenic plants were identified by PCR in grapefruit. RT-PCR showed that expression levels of#2and#17were higher than that in the WT. Treatment at4℃showed that#2and#17exhibited better cold tolerance. Conductivity measurement results showed that#2and#17was significantly lower than WT, indicating the degree of injury in transgenic plants lines were lower than that in the WT. DAB staining experiments showed that#2and#17dye depth lower than WT, indicating that the transgenic plants accumulated less H2O2.