Polyaniline/Attapugite/ PE(PAn-ATTP/PE)composites containing particles with core-shell structure were obtained via the two-step blending processs. The experimental condition is as follows: Organo-attapulgite and PAn w...Polyaniline/Attapugite/ PE(PAn-ATTP/PE)composites containing particles with core-shell structure were obtained via the two-step blending processs. The experimental condition is as follows: Organo-attapulgite and PAn was obtained by modifying attapulgite with laury benzenesulfonic acid sodium salt and, then added to PE. The electrical conductivity, structure and properties of the composites were studied. Under the function of shear stress, core-shell structure particles with ATTP as the core and PAn as the shell were formed in the composites. The structure of PAn-ATTP/PE composites were characterized by FTIR,XRD,SEM, etc, respectively. The effects of concentration of doping agent on the conductivity and mechanical property of the composites were investigated. The mechanical properties and impact fracture surface of the ternary composites were studied by means of the tensile tester, SEM, etc. The results show that polyaniline encapsulated ATTP enhances the strength of the PE. And the conductivity of PAn-ATTP/PE composites of is improved effectively when polyaniline encapsulated ATTP is added. The composite have good conductivity when 10% polyaniline encapsulated ATTP is added.展开更多
高效与特异的基因组定点修饰是基因工程动物研究的前沿与难点.链霉菌噬菌体ΦC31整合酶能介导含attB位点的外源基因定点整合于多种真核生物基因组的假attP位点,可维持外源基因的正常结构及高效表达.本文探讨ΦC31整合酶介导外源基因在...高效与特异的基因组定点修饰是基因工程动物研究的前沿与难点.链霉菌噬菌体ΦC31整合酶能介导含attB位点的外源基因定点整合于多种真核生物基因组的假attP位点,可维持外源基因的正常结构及高效表达.本文探讨ΦC31整合酶介导外源基因在猪基因组内定点整合的分子基础.构建含attB位点的报告载体pEGFP-N1-attB,与ΦC31整合酶表达载体pCMV-INT共转染猪肾PK15细胞,G418筛选获得单克隆细胞系.实时荧光定量PCR筛选出单拷贝整合的转基因细胞系.TAIL-PCR鉴定出1个猪基因组假attP位点,位于猪1号染色体,watson链,坐标114220087-114220126,命名为pig-attP-1.测序结果显示,pEGFP-N1-attB在attB位点处断开插入到pig-attP-1.用荧光计测定细胞培养基上清EGFP含量发现,该转基因细胞系EGFP的表达水平是本底的50倍(13500 AU vs.280 AU),表明pig-attP-1是利于外源基因高效表达的"友好位点".该研究不仅为实现外源基因在猪基因组内的定点整合提供了新策略,也为创制基因工程猪、建立动物生物反应器等研究注入了新思路.展开更多
Vibrio cholerae(V. cholerae) genome is equipped with a number of integrative mobile genetic element(IMGE) like prophages, plasmids, transposons or genomic islands, which provides fitness factors that help the pathogen...Vibrio cholerae(V. cholerae) genome is equipped with a number of integrative mobile genetic element(IMGE) like prophages, plasmids, transposons or genomic islands, which provides fitness factors that help the pathogen to survive in changing environmental conditions. Metagenomic analyses of clinical and environmental V. cholerae isolates revealed that dimer resolution sites(dif) harbor several structurally and functionally distinct IMGEs. All IMGEs present in the dif region exploit chromosomally encoded tyrosine recombinases, Xer C and Xer D, for integration. Integration takes place due to site-specific recombination between two specific DNA sequences; chromosomal sequence is called att B and IMGEs sequence is called att P. Different IMGEs present in the att P region have different attP structure but all of them are recognized by Xer C and Xer D enzymes and mediate either reversible or irreversible integration. Cholera toxin phage(CTXΦ), a lysogenic filamentous phage carrying the cholera toxin genes ctx AB, deserves special attention because it provides V. cholerae the crucial toxin and is always present in the dif region of all epidemic cholera isolates. Therefore, understanding the mechanisms of integration and dissemination of CTXΦ, genetic and ecological factors which support CTXΦ integration as well as production of virion from chromosomally integrated phage genome and interactions of CTXΦ with other genetic elements present in the genomes of V. cholerae is important for learning more about the biology of cholera pathogen.展开更多
A mercury biosensor was constructed by integrating biosensor genetic elements into E. coli JM109 chromosome in a single copy number, using the attP/attB recombination mechanism of λ phage. The genetic elements used i...A mercury biosensor was constructed by integrating biosensor genetic elements into E. coli JM109 chromosome in a single copy number, using the attP/attB recombination mechanism of λ phage. The genetic elements used include a regulatory protein gene （merR） along with operator/promoter （O/P） derived from the mercury resistance operon from pDU1358 plasmid of Serratia marcescens. The expression of reporter gene gfp is also controlled by merR/O/P. Integration of the construct into the chromosome was done to increase the stability and precision of the biosensor. This biosensor could detect Hg（Ⅱ） ions in the concentration range of 100–1700 nmol/L, and manifest the result as the expression of GFP. The GFP expression was significantly different （P 0.05） for each concentration of inducing Hg（Ⅱ） ions in the detection range, which reduces the chances of misinterpretation of results. A model using regression method was also derived for the quantification of the concentration of Hg（Ⅱ） in water samples.展开更多
The Streptomyces phage φC31 integrase can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes. To better understand the activity of φC31 integrase in the bovine genome...The Streptomyces phage φC31 integrase can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes. To better understand the activity of φC31 integrase in the bovine genome, DNA sequences of 44 integration events were analyzed, and 32 pseudo attP sites were identified. The majority of these sites share a sequence motif that contains inverted repeats and has similarities to wild-type attP site. Genomic DNA flanking these sites typically contained repetitive sequence elements, such as short and long interspersed repetitive elements. These sequence features indicate that DNA sequence recognition plays an important role in guiding φC31-mediated site-specific integration. In addition, BF27 integration hotspot sites were identified in the bovine genome, which accounted for 13.6% of all isolated integration events and mapped to an intron of the deleted in liver cancer 1 （DLC1） gene. Also we found that the pseudo attP sites in the bovine genome had other features in common with those in the human genome. This study represents the first time that the sequence features of pseudo attP sites specific integrase system has great potential for applied modifications in the bovine genome were analyzed. We conclude that this site- of the bovine genome.展开更多
文摘Polyaniline/Attapugite/ PE(PAn-ATTP/PE)composites containing particles with core-shell structure were obtained via the two-step blending processs. The experimental condition is as follows: Organo-attapulgite and PAn was obtained by modifying attapulgite with laury benzenesulfonic acid sodium salt and, then added to PE. The electrical conductivity, structure and properties of the composites were studied. Under the function of shear stress, core-shell structure particles with ATTP as the core and PAn as the shell were formed in the composites. The structure of PAn-ATTP/PE composites were characterized by FTIR,XRD,SEM, etc, respectively. The effects of concentration of doping agent on the conductivity and mechanical property of the composites were investigated. The mechanical properties and impact fracture surface of the ternary composites were studied by means of the tensile tester, SEM, etc. The results show that polyaniline encapsulated ATTP enhances the strength of the PE. And the conductivity of PAn-ATTP/PE composites of is improved effectively when polyaniline encapsulated ATTP is added. The composite have good conductivity when 10% polyaniline encapsulated ATTP is added.
文摘高效与特异的基因组定点修饰是基因工程动物研究的前沿与难点.链霉菌噬菌体ΦC31整合酶能介导含attB位点的外源基因定点整合于多种真核生物基因组的假attP位点,可维持外源基因的正常结构及高效表达.本文探讨ΦC31整合酶介导外源基因在猪基因组内定点整合的分子基础.构建含attB位点的报告载体pEGFP-N1-attB,与ΦC31整合酶表达载体pCMV-INT共转染猪肾PK15细胞,G418筛选获得单克隆细胞系.实时荧光定量PCR筛选出单拷贝整合的转基因细胞系.TAIL-PCR鉴定出1个猪基因组假attP位点,位于猪1号染色体,watson链,坐标114220087-114220126,命名为pig-attP-1.测序结果显示,pEGFP-N1-attB在attB位点处断开插入到pig-attP-1.用荧光计测定细胞培养基上清EGFP含量发现,该转基因细胞系EGFP的表达水平是本底的50倍(13500 AU vs.280 AU),表明pig-attP-1是利于外源基因高效表达的"友好位点".该研究不仅为实现外源基因在猪基因组内的定点整合提供了新策略,也为创制基因工程猪、建立动物生物反应器等研究注入了新思路.
基金Supported by Research in the Laboratory of Das B and NairGB is funded in part by Department of Science Technology,No.SB/FT/LS-309/2012Government of India(GOI)and the Department of Biotechnology,No.BT/MB/THSTI/HMC-SFC/2011Research in the Laboratory of Bhadra RK is partly financiallysupported by Council of Scientific and Industrial Research,GOIand Indian Council of Medical Research,GOI
文摘Vibrio cholerae(V. cholerae) genome is equipped with a number of integrative mobile genetic element(IMGE) like prophages, plasmids, transposons or genomic islands, which provides fitness factors that help the pathogen to survive in changing environmental conditions. Metagenomic analyses of clinical and environmental V. cholerae isolates revealed that dimer resolution sites(dif) harbor several structurally and functionally distinct IMGEs. All IMGEs present in the dif region exploit chromosomally encoded tyrosine recombinases, Xer C and Xer D, for integration. Integration takes place due to site-specific recombination between two specific DNA sequences; chromosomal sequence is called att B and IMGEs sequence is called att P. Different IMGEs present in the att P region have different attP structure but all of them are recognized by Xer C and Xer D enzymes and mediate either reversible or irreversible integration. Cholera toxin phage(CTXΦ), a lysogenic filamentous phage carrying the cholera toxin genes ctx AB, deserves special attention because it provides V. cholerae the crucial toxin and is always present in the dif region of all epidemic cholera isolates. Therefore, understanding the mechanisms of integration and dissemination of CTXΦ, genetic and ecological factors which support CTXΦ integration as well as production of virion from chromosomally integrated phage genome and interactions of CTXΦ with other genetic elements present in the genomes of V. cholerae is important for learning more about the biology of cholera pathogen.
基金Director, Central Institute of Fisheries Education, Mumbaifor providing facility and financial assistance in the form of Masters’ Fellowship during the research period
文摘A mercury biosensor was constructed by integrating biosensor genetic elements into E. coli JM109 chromosome in a single copy number, using the attP/attB recombination mechanism of λ phage. The genetic elements used include a regulatory protein gene （merR） along with operator/promoter （O/P） derived from the mercury resistance operon from pDU1358 plasmid of Serratia marcescens. The expression of reporter gene gfp is also controlled by merR/O/P. Integration of the construct into the chromosome was done to increase the stability and precision of the biosensor. This biosensor could detect Hg（Ⅱ） ions in the concentration range of 100–1700 nmol/L, and manifest the result as the expression of GFP. The GFP expression was significantly different （P 0.05） for each concentration of inducing Hg（Ⅱ） ions in the detection range, which reduces the chances of misinterpretation of results. A model using regression method was also derived for the quantification of the concentration of Hg（Ⅱ） in water samples.
基金supported by the grants from the National Science and Technology Major Project of China(Nos. 2009ZX08010-018B and 2011ZX08007-004)State & Shanghai Leading Academic Discipline(B204)
文摘The Streptomyces phage φC31 integrase can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes. To better understand the activity of φC31 integrase in the bovine genome, DNA sequences of 44 integration events were analyzed, and 32 pseudo attP sites were identified. The majority of these sites share a sequence motif that contains inverted repeats and has similarities to wild-type attP site. Genomic DNA flanking these sites typically contained repetitive sequence elements, such as short and long interspersed repetitive elements. These sequence features indicate that DNA sequence recognition plays an important role in guiding φC31-mediated site-specific integration. In addition, BF27 integration hotspot sites were identified in the bovine genome, which accounted for 13.6% of all isolated integration events and mapped to an intron of the deleted in liver cancer 1 （DLC1） gene. Also we found that the pseudo attP sites in the bovine genome had other features in common with those in the human genome. This study represents the first time that the sequence features of pseudo attP sites specific integrase system has great potential for applied modifications in the bovine genome were analyzed. We conclude that this site- of the bovine genome.
