Sequence analysis of a new norovirus(NV) isolated from Lanzou city of China was performed based on partial sequence of RNA dependent RNA polymerase(RdRp) and complete capsid protein(VP1) gene.The isolated strain CHN02...Sequence analysis of a new norovirus(NV) isolated from Lanzou city of China was performed based on partial sequence of RNA dependent RNA polymerase(RdRp) and complete capsid protein(VP1) gene.The isolated strain CHN02/LZ35666 shared high sequence homology with GII-4 NVs.Nucleotide homologies of RdRp region and encoded capsid protein region were 90.4%-98.6% and 89.8%-95.7%,respectively,while amino acid homology of capsid protein region was 94.4%-97.4%.The analysis of GDD motif in RdRp region indicated this GDD motif of Lanzhou strain differed from those of the GII-4 predominant epidemic strains.Lanzhou strain formed an independent branch in GII-4 cluster in the phylogenetic tree based on nucleotide sequence of RdRp region and amino acid sequence of capsid protein.Sequence alignment revealed a mutation at the fourth key site of the receptor-binding interface in the strains isolated after 2002 compared with those of previous strains suggesting a possible change of binding pattern to HBGAs receptors.展开更多
The RNA-dependent RNA polymerase (RdRp) of SARS-Coronavirus plays a crucial action in the processes of viral replication and discontinuous transcription.Because of its important function and its highest conservation a...The RNA-dependent RNA polymerase (RdRp) of SARS-Coronavirus plays a crucial action in the processes of viral replication and discontinuous transcription.Because of its important function and its highest conservation among the Coronaviruses,SARS-RdRp will be a potential drug target for anti-SARS therapy.Here we map the conserved motifs in SARS-RdRp by multiple sequence alignment,combined with structural information,in groups of virus RdRps proteins which are evolutionarily related,but share significantly low sequence similarity(10%-20%).Besides six known motifs,three novel ones are identified.Secondary structure and three dimensional structure indicate that conserved motifs tend to be situated in the junction regions of conserved beta sheets or alpha helices,whose location and length are conserved,but the primary amino acid sequences do not display conservation.SARS-RdRp shows similar structure to other viral RdRps and this implies that the current available antiviral agents for other RdRps might have important implications for anti-SARS therapy.展开更多
Fluorinated polymers are receiving more and more attention worldwide due to their unique chemical properties,and modified fluorinated polymers with different topologies are persued for enriching and enhancing their pe...Fluorinated polymers are receiving more and more attention worldwide due to their unique chemical properties,and modified fluorinated polymers with different topologies are persued for enriching and enhancing their performance in a variety of application fields.In this work,main-chain-type semifluorinated graft copolymers are produced steadily in continuous tube reactors via photocontrolled step transferaddition and radical-termination(START)polymerization and Cu(0)-mediated reversible deactivation radical polymerization(Cu(0)-RDRP)at room temperature for the first time.Specifically,semifluorinated alternating copolymer(AB)n B is prepared by START polymerization of 1,6-diiodoperfluorohexane(A)and 1,7-octadiene(B)in the first quartz pipeline under irradiation with purple LED light at 20℃.The(AB)nB with periodic C―I bonds is then flowed into the second copper pipeline directly and acts as the macroinitiators for Cu(0)-RDRP of methyl acrylate(MA)to obtain corresponding graft copolymer(AB)n B-g-PMA.This work provides a new strategy for continuous synthesis of fluorinated graft copolymer materials.展开更多
目的构建戊型肝炎病毒(Hepatitis E virus,HEV)RdRp基因真核表达质粒,并检测其在PLC/PRF/5、A549和HepG2 3种细胞中的表达,为后续RdRp的研究提供实验依据。方法利用RT-nPCR法从阳性HEV粪便中扩增RdRp基因片段,插入pcDNA3.0真核表达载体...目的构建戊型肝炎病毒(Hepatitis E virus,HEV)RdRp基因真核表达质粒,并检测其在PLC/PRF/5、A549和HepG2 3种细胞中的表达,为后续RdRp的研究提供实验依据。方法利用RT-nPCR法从阳性HEV粪便中扩增RdRp基因片段,插入pcDNA3.0真核表达载体中,并在其3′端插入EGFP报告基因,构建融合表达质粒pcDNA3.0-RdRp-EGFP,脂质体法转染PLC/PRF/5、A549和HepG2细胞,荧光显微镜观察报告基因的表达,RT-nPCR检测RdRp基因mRNA的转录情况,Westernblot检测RdRp蛋白的表达。结果重组表达质粒经酶切鉴定和测序证实构建正确;RdRp基因和蛋白在A549和HepG2细胞中可高效、特异性表达。结论成功构建了RdRp基因真核表达质粒,并在A549和HepG2培养细胞中大量表达,为进一步研究RdRp在HEV复制过程中的功能奠定了基础。展开更多
文摘Sequence analysis of a new norovirus(NV) isolated from Lanzou city of China was performed based on partial sequence of RNA dependent RNA polymerase(RdRp) and complete capsid protein(VP1) gene.The isolated strain CHN02/LZ35666 shared high sequence homology with GII-4 NVs.Nucleotide homologies of RdRp region and encoded capsid protein region were 90.4%-98.6% and 89.8%-95.7%,respectively,while amino acid homology of capsid protein region was 94.4%-97.4%.The analysis of GDD motif in RdRp region indicated this GDD motif of Lanzhou strain differed from those of the GII-4 predominant epidemic strains.Lanzhou strain formed an independent branch in GII-4 cluster in the phylogenetic tree based on nucleotide sequence of RdRp region and amino acid sequence of capsid protein.Sequence alignment revealed a mutation at the fourth key site of the receptor-binding interface in the strains isolated after 2002 compared with those of previous strains suggesting a possible change of binding pattern to HBGAs receptors.
文摘The RNA-dependent RNA polymerase (RdRp) of SARS-Coronavirus plays a crucial action in the processes of viral replication and discontinuous transcription.Because of its important function and its highest conservation among the Coronaviruses,SARS-RdRp will be a potential drug target for anti-SARS therapy.Here we map the conserved motifs in SARS-RdRp by multiple sequence alignment,combined with structural information,in groups of virus RdRps proteins which are evolutionarily related,but share significantly low sequence similarity(10%-20%).Besides six known motifs,three novel ones are identified.Secondary structure and three dimensional structure indicate that conserved motifs tend to be situated in the junction regions of conserved beta sheets or alpha helices,whose location and length are conserved,but the primary amino acid sequences do not display conservation.SARS-RdRp shows similar structure to other viral RdRps and this implies that the current available antiviral agents for other RdRps might have important implications for anti-SARS therapy.
基金financially supported by the National Natural Science Foundation of China(Nos.22071168,21971178 and 21674071)the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD)。
文摘Fluorinated polymers are receiving more and more attention worldwide due to their unique chemical properties,and modified fluorinated polymers with different topologies are persued for enriching and enhancing their performance in a variety of application fields.In this work,main-chain-type semifluorinated graft copolymers are produced steadily in continuous tube reactors via photocontrolled step transferaddition and radical-termination(START)polymerization and Cu(0)-mediated reversible deactivation radical polymerization(Cu(0)-RDRP)at room temperature for the first time.Specifically,semifluorinated alternating copolymer(AB)n B is prepared by START polymerization of 1,6-diiodoperfluorohexane(A)and 1,7-octadiene(B)in the first quartz pipeline under irradiation with purple LED light at 20℃.The(AB)nB with periodic C―I bonds is then flowed into the second copper pipeline directly and acts as the macroinitiators for Cu(0)-RDRP of methyl acrylate(MA)to obtain corresponding graft copolymer(AB)n B-g-PMA.This work provides a new strategy for continuous synthesis of fluorinated graft copolymer materials.
文摘目的构建戊型肝炎病毒(Hepatitis E virus,HEV)RdRp基因真核表达质粒,并检测其在PLC/PRF/5、A549和HepG2 3种细胞中的表达,为后续RdRp的研究提供实验依据。方法利用RT-nPCR法从阳性HEV粪便中扩增RdRp基因片段,插入pcDNA3.0真核表达载体中,并在其3′端插入EGFP报告基因,构建融合表达质粒pcDNA3.0-RdRp-EGFP,脂质体法转染PLC/PRF/5、A549和HepG2细胞,荧光显微镜观察报告基因的表达,RT-nPCR检测RdRp基因mRNA的转录情况,Westernblot检测RdRp蛋白的表达。结果重组表达质粒经酶切鉴定和测序证实构建正确;RdRp基因和蛋白在A549和HepG2细胞中可高效、特异性表达。结论成功构建了RdRp基因真核表达质粒,并在A549和HepG2培养细胞中大量表达,为进一步研究RdRp在HEV复制过程中的功能奠定了基础。
