CRISPR/Cas9[Clustered regular interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9]系统是广泛存在于细菌和古生菌中可降解入侵病毒和噬菌体DNA的一种基因定点编辑技术,由于其具有快速、精准、高效,易于设计...CRISPR/Cas9[Clustered regular interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9]系统是广泛存在于细菌和古生菌中可降解入侵病毒和噬菌体DNA的一种基因定点编辑技术,由于其具有快速、精准、高效,易于设计,且特异性高等优势,得到了广泛的应用。概述了CRISPR/Cas9基因编辑技术的发展历程,并介绍其结构和编辑原理、在动物细胞系构建中的应用及展望,以便为更合理地应用和改进该技术提供系统的文献参考。展开更多
We have constructed a retroviral vector mediated mammalian cell expression system of the capsid precursor protein of foot-and-mouth disease virus(FMDV).The recombinant retroviral vector pBABEpuro-P1-2A-EGFP was constr...We have constructed a retroviral vector mediated mammalian cell expression system of the capsid precursor protein of foot-and-mouth disease virus(FMDV).The recombinant retroviral vector pBABEpuro-P1-2A-EGFP was constructed by sequentially inserting capsid precursor protein gene(P1) of FMDV and enhanced green fluorescent protein gene(EGFP) into pBABEpuro.The recombinant retroviral vector and the pVSV-G plasmid were co-transfected into packaging cells(GP2-293) by liposomemediated transduction to produce the pseudovirus.The pseudovirus was used to infect BHK-21 cells and resistant cells were screened with puromycin.Green fluorescent proteins were observed by fluorescence microscopy and expression of the capsid precursor protein gene of FMDV was detected by indirect immunofluorescence.The recombinant retroviral vector pBABEpuro-P1-2A-EGFP was constructed successfully.The capsid precursor protein of FMDV and green fluorescent protein were expressed in BHK-21 cells.The mammalian cell expression system for the capsid precursor protein of FMDV has been constructed successfully,which lays the foundation of development of a FMDV subunit vaccine.展开更多
文摘CRISPR/Cas9[Clustered regular interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9]系统是广泛存在于细菌和古生菌中可降解入侵病毒和噬菌体DNA的一种基因定点编辑技术,由于其具有快速、精准、高效,易于设计,且特异性高等优势,得到了广泛的应用。概述了CRISPR/Cas9基因编辑技术的发展历程,并介绍其结构和编辑原理、在动物细胞系构建中的应用及展望,以便为更合理地应用和改进该技术提供系统的文献参考。
基金supported by the National Programs for High Technology Research and Development of China (2006AA10A204)the Gansu Key Technologies R&D Program(ZGS-052-A41-0006-03)the Programs for Director Fund of Lanzhou Veterinary Research Institute
文摘We have constructed a retroviral vector mediated mammalian cell expression system of the capsid precursor protein of foot-and-mouth disease virus(FMDV).The recombinant retroviral vector pBABEpuro-P1-2A-EGFP was constructed by sequentially inserting capsid precursor protein gene(P1) of FMDV and enhanced green fluorescent protein gene(EGFP) into pBABEpuro.The recombinant retroviral vector and the pVSV-G plasmid were co-transfected into packaging cells(GP2-293) by liposomemediated transduction to produce the pseudovirus.The pseudovirus was used to infect BHK-21 cells and resistant cells were screened with puromycin.Green fluorescent proteins were observed by fluorescence microscopy and expression of the capsid precursor protein gene of FMDV was detected by indirect immunofluorescence.The recombinant retroviral vector pBABEpuro-P1-2A-EGFP was constructed successfully.The capsid precursor protein of FMDV and green fluorescent protein were expressed in BHK-21 cells.The mammalian cell expression system for the capsid precursor protein of FMDV has been constructed successfully,which lays the foundation of development of a FMDV subunit vaccine.
