[Objective] The paper aimed to construct a new T vector based on plasmid pUC19 digested with Xcm Ⅰ. [Method] Two complementary oligonucleotide chains containing two Xcm Ⅰ sites were synthesized. After denaturation a...[Objective] The paper aimed to construct a new T vector based on plasmid pUC19 digested with Xcm Ⅰ. [Method] Two complementary oligonucleotide chains containing two Xcm Ⅰ sites were synthesized. After denaturation and renaturation,the adaptor was cloned into plasmid pUC19 between the Hind Ⅲ and BamH Ⅰ sites. The new plasmid,pUC19-HB-T vector,was digested with Xcm Ⅰ to derive a T-vector with 3′ end overhanging a T base. [Result] The constructed pUC19-HB-T vector was efficient in cloning PCR products,with an efficiency of 95% at least. [Conclusion] A new Xcm Ⅰ-based pUC19-HB-T vector was constructed,which could be applied to cloning of PCR products and other microbiology operations.展开更多
[Objective] The aim was to construct a recombinant pseudorabies virus expressing canine distemper virus H gene and investigate its biological characters.[Method] H gene of canine distemper virus（CDV）strain Onderstep...[Objective] The aim was to construct a recombinant pseudorabies virus expressing canine distemper virus H gene and investigate its biological characters.[Method] H gene of canine distemper virus（CDV）strain Onderstepoort was produced by RT-PCR,inserted into pcDNA3.1（＋）vector to construct a expression cassette,which was then subcloned into transfer vector p8AA,prior to the insertion of LacZ expression cassette.The resulting new transfer vector was named as p8AAZH.Subsequently,p8AAZH was co-transfected with the genome of pseudorabies virus（PRV）Bartha-K61 into BHK-21 cells to enable gene recombination and virus package,and the virus solution was collected as cytopathic effect occurring.A series of procedures including blue plaque purification,PCR identification,observation under electron microscope and Western blot were carried out to screen the recombinant pseudorabies virus and identify the protein expression of target gene.Meanwhile,growth curve of the recombinant virus was determined in BHK-21 cells.[Result] The H gene had been inserted into the genome of Bartha-K61 strain,and RPRV-H was the same as Bartha-K61 in the one-step growth curve and cytopathic effect in BHK-21 cells.[Conclusion] The recombinant pseudorabies virus was constructed,and the insertion of H gene did not influence proliferation of recombinant virus,which laid a foundation for development of recombinant canine distemper virus vaccine.展开更多
随着油耗法规和排放法规的日趋严格,市场对高效节能内燃机的需求越来越高,GDI汽油机独特的油气混合方式,热效率明显高于传统PFI汽油机,GDI汽油机逐渐成为汽车内燃机的开发主流之一。GDI汽油机在性能测试过程中,同一台发动机相同的测试设...随着油耗法规和排放法规的日趋严格,市场对高效节能内燃机的需求越来越高,GDI汽油机独特的油气混合方式,热效率明显高于传统PFI汽油机,GDI汽油机逐渐成为汽车内燃机的开发主流之一。GDI汽油机在性能测试过程中,同一台发动机相同的测试设备,不同的进气湿度下,外特性有明显差异,本文选择一台2.0 L GDI汽油机进行不同湿度的外特性试验,探究湿度对GDI汽油机外特性的影响。展开更多
随着乘用车小排量增压发动机应用的增多,增压发动机的早燃问题也越来越常见,通过排查一台1.4 T PFI增压汽油机在整车试验中出现的连杆断裂问题,验证了低水温条件下PFI增压汽油机会发生早燃,并对早燃识别的水温阈值进行优化,通过转速和...随着乘用车小排量增压发动机应用的增多,增压发动机的早燃问题也越来越常见,通过排查一台1.4 T PFI增压汽油机在整车试验中出现的连杆断裂问题,验证了低水温条件下PFI增压汽油机会发生早燃,并对早燃识别的水温阈值进行优化,通过转速和出水温度的扫点验证,得到低水温条件下早燃的发生规律。展开更多
文摘[Objective] The paper aimed to construct a new T vector based on plasmid pUC19 digested with Xcm Ⅰ. [Method] Two complementary oligonucleotide chains containing two Xcm Ⅰ sites were synthesized. After denaturation and renaturation,the adaptor was cloned into plasmid pUC19 between the Hind Ⅲ and BamH Ⅰ sites. The new plasmid,pUC19-HB-T vector,was digested with Xcm Ⅰ to derive a T-vector with 3′ end overhanging a T base. [Result] The constructed pUC19-HB-T vector was efficient in cloning PCR products,with an efficiency of 95% at least. [Conclusion] A new Xcm Ⅰ-based pUC19-HB-T vector was constructed,which could be applied to cloning of PCR products and other microbiology operations.
文摘[Objective] The aim was to construct a recombinant pseudorabies virus expressing canine distemper virus H gene and investigate its biological characters.[Method] H gene of canine distemper virus（CDV）strain Onderstepoort was produced by RT-PCR,inserted into pcDNA3.1（＋）vector to construct a expression cassette,which was then subcloned into transfer vector p8AA,prior to the insertion of LacZ expression cassette.The resulting new transfer vector was named as p8AAZH.Subsequently,p8AAZH was co-transfected with the genome of pseudorabies virus（PRV）Bartha-K61 into BHK-21 cells to enable gene recombination and virus package,and the virus solution was collected as cytopathic effect occurring.A series of procedures including blue plaque purification,PCR identification,observation under electron microscope and Western blot were carried out to screen the recombinant pseudorabies virus and identify the protein expression of target gene.Meanwhile,growth curve of the recombinant virus was determined in BHK-21 cells.[Result] The H gene had been inserted into the genome of Bartha-K61 strain,and RPRV-H was the same as Bartha-K61 in the one-step growth curve and cytopathic effect in BHK-21 cells.[Conclusion] The recombinant pseudorabies virus was constructed,and the insertion of H gene did not influence proliferation of recombinant virus,which laid a foundation for development of recombinant canine distemper virus vaccine.
文摘随着油耗法规和排放法规的日趋严格,市场对高效节能内燃机的需求越来越高,GDI汽油机独特的油气混合方式,热效率明显高于传统PFI汽油机,GDI汽油机逐渐成为汽车内燃机的开发主流之一。GDI汽油机在性能测试过程中,同一台发动机相同的测试设备,不同的进气湿度下,外特性有明显差异,本文选择一台2.0 L GDI汽油机进行不同湿度的外特性试验,探究湿度对GDI汽油机外特性的影响。
