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Objective To construct active recombinant caspases 3 gene(r caspases 3)eukaryotic expression plasmid and observe the apoptosis inducing activity of r caspase 3 in pancreatic carcinoma cells. Methods pcDNA3.1(+)/r caspase 3 was constructed and pancreatic carcinoma cells(PC Ⅱ)were transfected with the pcDNA3.1(+)/r caspases 3 by liposomes(LipofectAMINE).The expression of r Caspase 3 mRNA in pancreatic carcinoma cells was detected by reverse transcription process of the polymerase chain reaction(RT PCR), and the signs of apoptosis were examined in pancreatic carcinoma cells by the methods of the DNA electrophoresis and flow cytometry analysis(FACS).Results The sequence inserted in pBlueSKM/r Caspase 3 plasmid was coincident with that of the r caspases 3. The evaluation result of pcDNA3.1(+)/r caspases 3 through enzyme cutting was correct. A 894bp strap was observed by RT PCR after pancreatic carcinoma cells being transfected with the pcDNA3.1(+)/r caspases 3 by liposomes. No strap was found in control groups. A characteristic DNA ladder was observed in pancreatic carcinoma cells DNA electrophoresis, and transparent hypodiploid karyotype peak was found by FACS. Conclusion The plasmid of pcDNA3.1(+)/r Caspase 3 was constructed successfully, the expression of r Caspase 3 mRMA in pancreatic carcinoma cells was confirmed by RT PCR, and pcDNA3.1(+)/r Caspase 3 can induce apoptosis in pancreatic carcinoma cells.
Objective To construct active recombinant caspases 3 gene(r caspases 3)eukaryotic expression plasmid and observe the apoptosis inducing activity of r caspase 3 in pancreatic carcinoma cells. Methods pcDNA3.1(+)/r caspase 3 was constructed and pancreatic carcinoma cells(PC II)were transfected with the pcDNA3.1(+)/r caspases 3 by liposomes(LipofectAMINE). The expression of r Caspase 3 mRNA in pancreatic carcinoma cells was detected by reverse transcription process of the polymerase chain reaction (RT PCR), and The signs of apoptosis were examined in pancreatic carcinoma cells by the methods of the DNA electrophoresis and flow cytometry analysis(FACS).Results The sequence inserted in pBlueSKM/r Caspase 3 plasmid was coincident with that of the r caspases 3. The evaluation result of pcDNA3.1(+)/r caspases 3 through enzyme cutting was correct. A 894bp strap was observed by RT PCR after pancreatic carcinoma cells are transfected with the pcDNA3.1(+)/r caspases 3 by a characteristic DNA ladder was observed in pancreatic carcinoma cells DNA electrophoresis, and transparent hypodiploid karyotype peak was found by FACS. Conclusion The plasmid of pcDNA3.1(+)/r Caspase 3 was used successfully. , the expression of r Caspase 3 mRMA in pancreatic carcinoma cells was confirmed by RT PCR, and pcDNA3.1(+)/r Caspase 3 can induce apoptosis in pancreatic carcinoma cells.