Ojective: To investigate the effect of hammerhead ribozyme that specifically cleaves c-myc mRNA on the proliferation of vascular smooth muscle cells (VSMCs ). Methods: Based on the computer analysis of the secondary s...Ojective: To investigate the effect of hammerhead ribozyme that specifically cleaves c-myc mRNA on the proliferation of vascular smooth muscle cells (VSMCs ). Methods: Based on the computer analysis of the secondary structure of c-myc mRNA, nt 2029 in rat c-myc oncogene was selected as a cleaving site for hammerhead ribozyme and the ribozyme was designed. With automatic DNA synthesizer, the two complementary DNA strands of the ribozyme were synthesized. The ribozyme gene was cloned into pGEM3Zf ( + ) vector and subcloned into eukaryotic expression pcD-NA3 vector. The recombinant pcDNA-Rz was transfected into the cultured rat VSMCs by lipofectAMINE mediated DNA transfection protocol and individual cell clones were selected by G418. Results: The sequence of ribozyme gene inserted in pGEMSZf ( + ) vector was proved to be perfectly correct. In VSMCs transfected with recombinant pcDNA-Rz, flow cytometry analysis showed that the S phase and G2/M fractions were decreased significantly and cell proliferation stagnated in the G0/G1 phase. Conclusion: The results suggest that hammerhead ribozyme that specifically cleaves c-myc mRNA can significantly inhibit the proliferation of VSMCs.展开更多
A label-free colorimetric protocol based on peptide nucleic acid/silver nanoparticles(PNA/Ag NPs) has been initially proposed for specific recognition of m RNA.Making use of the controlled silver nanoparticles aggrega...A label-free colorimetric protocol based on peptide nucleic acid/silver nanoparticles(PNA/Ag NPs) has been initially proposed for specific recognition of m RNA.Making use of the controlled silver nanoparticles aggregation/dispersion by PNA/PNA–RNA complex, proto-oncogene c-Myc m RNA detection can be achieved. Moreover, the PNA/Ag NPs platform can undergo color change in response to target c-Myc m RNA with single-base-mismatch sensitivity, which could further help in visually identify single nucleotide differences in target genes.展开更多
基金Supported by the National Natural Science Foundation of China(No.39600064)
文摘Ojective: To investigate the effect of hammerhead ribozyme that specifically cleaves c-myc mRNA on the proliferation of vascular smooth muscle cells (VSMCs ). Methods: Based on the computer analysis of the secondary structure of c-myc mRNA, nt 2029 in rat c-myc oncogene was selected as a cleaving site for hammerhead ribozyme and the ribozyme was designed. With automatic DNA synthesizer, the two complementary DNA strands of the ribozyme were synthesized. The ribozyme gene was cloned into pGEM3Zf ( + ) vector and subcloned into eukaryotic expression pcD-NA3 vector. The recombinant pcDNA-Rz was transfected into the cultured rat VSMCs by lipofectAMINE mediated DNA transfection protocol and individual cell clones were selected by G418. Results: The sequence of ribozyme gene inserted in pGEMSZf ( + ) vector was proved to be perfectly correct. In VSMCs transfected with recombinant pcDNA-Rz, flow cytometry analysis showed that the S phase and G2/M fractions were decreased significantly and cell proliferation stagnated in the G0/G1 phase. Conclusion: The results suggest that hammerhead ribozyme that specifically cleaves c-myc mRNA can significantly inhibit the proliferation of VSMCs.
基金the National Natural Science Foundation of China (21305058, 21205056, 21075058 and 21503104)Tai-Shan Scholar Research Fund of Shandong Province
文摘A label-free colorimetric protocol based on peptide nucleic acid/silver nanoparticles(PNA/Ag NPs) has been initially proposed for specific recognition of m RNA.Making use of the controlled silver nanoparticles aggregation/dispersion by PNA/PNA–RNA complex, proto-oncogene c-Myc m RNA detection can be achieved. Moreover, the PNA/Ag NPs platform can undergo color change in response to target c-Myc m RNA with single-base-mismatch sensitivity, which could further help in visually identify single nucleotide differences in target genes.
