| The abitotic stresses serve as the important environmental factors that influence the normal growth of plants.Among these factors,the stress of low temperature is one of the most harmful factors.The low temperature can cause the damage in different degrees, even more death in plants. Plants develop a variety of sophisticated mechanisms,which including the effect of low temperature response genes and the signal pathway,to response the changing environment and protect themselves from environmental stress.In this study,we uesd Chorispora Bungeana which is a rare alpine subnival plant species as material. The technology of genetically modified and virus-induced gene silencing(VIGS) were used to verify the functionalities of CbPLDα, CbPLDβ,CbPLDδ gene from transgenic tobacco,and analysis the response mechanisms under low temperature stress.The main results are as follows:(1) Introduced CbPLDδ into tobacco with the CaMV 35 S promoter. The transgenic tobacco plants were detected by PCR.The sequences results showed that the CbPLDδ genes have been transferred into tobacco successfully.(2) Constructed a TRV-CbPLDα,TRV-CbPLDβ and TRV-CbPLDδ virul vectors and introduced into tobacco by Agrobacterium-mediated injection.The RT-PCR were used to detect the gene silencing rate of CbPLDα,CbPLDβ and CbPLDδ in transgenic tobacco.Then,we filtered the highest rate of gene silencing tobacco and named as TRV-CbPLDα1,TRV-CbPLDβ 3 and TRV-CbPLDδ2,respectively.(3) The expression levels of CbPLDα, CbPLDβ and CbPLDδ gene in transgenic tobacco under low temperature stress(4℃) were studied.The result showed that,low temperature induced the expression of CbPLDα,CbPLDβ and CbPLDδ,and the expression of CbPLDβ gene behind the CbPLDα and CbPLDδ with the time prolong under low temperature stress.(4) The metabolism regulations of CbPLD transgenic tobaccos and TRV-CbPLDgene silence tobaccos in response to low temperature stress were analyzed. The results showed that both CbPLD transgenic tobaccos and TRV-CbPLD gene silence tobaccos involved in the regulation pathway of membrane stability, osmotic adjustment and the regulation of anti-oxidative system. CbPLDα, CbPLDβ and CbPLDδ gene can provide powerful genetic resources for improving low temperature resistance of plants. |
