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
Dunaliella salina is known as one of the most salt-tolerant eukaryotic or- ganisms, and the most ideal model organism for studying plant adaption to high salinity. In recent years, the study on molecular mechanism of ...Dunaliella salina is known as one of the most salt-tolerant eukaryotic or- ganisms, and the most ideal model organism for studying plant adaption to high salinity. In recent years, the study on molecular mechanism of salt tolerance in Dunaliella salina has become the focus of scholars at home and abroad with the development of molecular biological techniques. This study reviewed studies on adaption of Dunaliella salina to high salinity in aspects of osmotic adjustment, salt tolerance-related genes and proteins of Dunaliella salina and signal transduction pathway of salt stress.展开更多
Dunaliella salina is an important model organism for investigating the salt tolerance mechanism of plant. MAPK is the key gene in the molecular pathway of salt tolerance of plant. In this experiment, the open reading ...Dunaliella salina is an important model organism for investigating the salt tolerance mechanism of plant. MAPK is the key gene in the molecular pathway of salt tolerance of plant. In this experiment, the open reading frame （ORF） of DsMAPK gene was amplified by PCR. The target fragment was cloned in pGS-21a, a prokaryotic expression vector with GST-tag. The recombinant plasmid pGS-21a- DsMAPK was then transformed into E. coil BL21 （DE3）. The expression was induced with IPTG. Then the expression form of the recombinant protein was analyzed. The expression products were purified with GST-SefinoseTM Kit and identified with SDS-PAGE and Western blot. The results showed the recombinant expression vector pGS-21a-DsMAPK was constructed successfully, and the molecular weight of the expressed recombinant protein was as same as expected. Western blot analysis showed the purified recombinant protein could be identified specially by the anti- GST antibody and had a good immunological activity. The successful expression of DsMAPK will lay a basis for the further research on the role of DsMAPK in the salt tolerance mechanism at the protein level.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(3147226030972240)~~
文摘Dunaliella salina is known as one of the most salt-tolerant eukaryotic or- ganisms, and the most ideal model organism for studying plant adaption to high salinity. In recent years, the study on molecular mechanism of salt tolerance in Dunaliella salina has become the focus of scholars at home and abroad with the development of molecular biological techniques. This study reviewed studies on adaption of Dunaliella salina to high salinity in aspects of osmotic adjustment, salt tolerance-related genes and proteins of Dunaliella salina and signal transduction pathway of salt stress.
基金Supported by National Natural Science Foundation of China(3147226030972240)~~
文摘Dunaliella salina is an important model organism for investigating the salt tolerance mechanism of plant. MAPK is the key gene in the molecular pathway of salt tolerance of plant. In this experiment, the open reading frame （ORF） of DsMAPK gene was amplified by PCR. The target fragment was cloned in pGS-21a, a prokaryotic expression vector with GST-tag. The recombinant plasmid pGS-21a- DsMAPK was then transformed into E. coil BL21 （DE3）. The expression was induced with IPTG. Then the expression form of the recombinant protein was analyzed. The expression products were purified with GST-SefinoseTM Kit and identified with SDS-PAGE and Western blot. The results showed the recombinant expression vector pGS-21a-DsMAPK was constructed successfully, and the molecular weight of the expressed recombinant protein was as same as expected. Western blot analysis showed the purified recombinant protein could be identified specially by the anti- GST antibody and had a good immunological activity. The successful expression of DsMAPK will lay a basis for the further research on the role of DsMAPK in the salt tolerance mechanism at the protein level.
