The exogenous gene was integrated into Dunaliella salina successfully by using LiAc/PEG mediating method for the first time. According to the results of histochemical staining, transgenic D. salina was blue, showing t...The exogenous gene was integrated into Dunaliella salina successfully by using LiAc/PEG mediating method for the first time. According to the results of histochemical staining, transgenic D. salina was blue, showing that the exogenous GUS gene was successfully expressed in the cells of D. salina. Meanwhile, the effects of growth state of D. salina, plasmid concentration and temperature on its transformation efficiency were studied, and the transformation conditions were optimized. The results show that the optimum conditions for the genetic transformation of D. salina are shown as follows： D. salina was in the early logarithmic phase; plasmid DNA concentration was 600 μg/ml; temperature was 29 ℃, and transformation efficiency was up to 74.8‰ under the best conditions. According to the results of PCR amplification and PCR-Southern hybridization, the target gene had been integrated into genome and was hereditary.展开更多
[Objectives] This study was conducted to investigate the function of Dunaliella salina calmodulin kinase(CaM K) gene.[Methods] The sense and antisense gene fragments(223 bp) and spacer sequence(129 bp) of D.salina cal...[Objectives] This study was conducted to investigate the function of Dunaliella salina calmodulin kinase(CaM K) gene.[Methods] The sense and antisense gene fragments(223 bp) and spacer sequence(129 bp) of D.salina calmodulin kinase gene were cloned and inserted into the downstream part of the35 S promoter of the eukaryotic expression vector pM DCMGN-Cat.The siRNA expression system of CaM K gene was successfully constructed.The p CaM K-RNAi expression vector was transformed into D.salina cells by the LiA c/PEG-mediated method,giving transgenic D.salina.The expression of CaM K gene was then analyzed by real-time fluorescence quantitative PCR.[Results]The expression of CaM K gene in the transgenic D.salina was significantly reduced,by 70% compared with the control group,suggesting that the expression of CaM K gene was significantly inhibited.The examination of the growth status of D.salina showed that D.salina cell division and proliferation were also affected.It is proved that CaM K gene has a positive regulation effect on the division and proliferation of D.salina cells.[Conclusions] The study provides important information for further elucidating the function and action mechanism of D.salina calmodulin kinase gene.展开更多
基金Supported by National Natural Science Foundation of China(31472260)~~
文摘The exogenous gene was integrated into Dunaliella salina successfully by using LiAc/PEG mediating method for the first time. According to the results of histochemical staining, transgenic D. salina was blue, showing that the exogenous GUS gene was successfully expressed in the cells of D. salina. Meanwhile, the effects of growth state of D. salina, plasmid concentration and temperature on its transformation efficiency were studied, and the transformation conditions were optimized. The results show that the optimum conditions for the genetic transformation of D. salina are shown as follows： D. salina was in the early logarithmic phase; plasmid DNA concentration was 600 μg/ml; temperature was 29 ℃, and transformation efficiency was up to 74.8‰ under the best conditions. According to the results of PCR amplification and PCR-Southern hybridization, the target gene had been integrated into genome and was hereditary.
基金Supported by National Natural Science Foundation of China(31472260,30972240)。
文摘[Objectives] This study was conducted to investigate the function of Dunaliella salina calmodulin kinase(CaM K) gene.[Methods] The sense and antisense gene fragments(223 bp) and spacer sequence(129 bp) of D.salina calmodulin kinase gene were cloned and inserted into the downstream part of the35 S promoter of the eukaryotic expression vector pM DCMGN-Cat.The siRNA expression system of CaM K gene was successfully constructed.The p CaM K-RNAi expression vector was transformed into D.salina cells by the LiA c/PEG-mediated method,giving transgenic D.salina.The expression of CaM K gene was then analyzed by real-time fluorescence quantitative PCR.[Results]The expression of CaM K gene in the transgenic D.salina was significantly reduced,by 70% compared with the control group,suggesting that the expression of CaM K gene was significantly inhibited.The examination of the growth status of D.salina showed that D.salina cell division and proliferation were also affected.It is proved that CaM K gene has a positive regulation effect on the division and proliferation of D.salina cells.[Conclusions] The study provides important information for further elucidating the function and action mechanism of D.salina calmodulin kinase gene.
