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Highly Sensitive Fluorescent-labeled Probes and Glass Slide Hybridization for the Detection of Plant RNA Viruses and a Viroid

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Acta Biochimica et Biophysica Sinica 2007年第5期39卷 326-334页

作者： Zhiyou DU~(1,2), Bo JIN~3, Wenhong LIU~4, Liang CHEN~1, and Jishuang CHEN~(2,1*) 1 College of Life sciences, zhejiang university, hangzhou 310029, china 2 institute of bioengineering, zhejiang sci-tech university, hangzhou 310018, china 3 College of Life sciences, zhejiang Chinese Medical university, hangzhou 310053, china 4 College of bioengineering, zhejiang Chinese Medical university, hangzhou 310053, china College of Life sciences Zhejiang UniversityHangzhou 310029 China institute of bioengineering Zhejiang Sci-Tech UniversityHangzhou 310018 China College of Life sciences Zhejiang Chinese Medical UniversityHangzhou 310053 China College of bioengineering Zhejiang Chinese Medical UniversityHangzhou 310053 China

In this study, a modified method of the conventional RNA dot-blot hybridization was established, by replacing32p labels with CY5 labels and replacing nylon membranes with positive-charged glass slides, for detecting plant RNA viruses and a viroid. The modified RNA dot-blot hybridization method was named glass slide hybridization. The optimum efficiency of RNA binding onto the surfaces of activated glass slide was achieved using aminosilane-coated glass slide as a solid matrix and 5×saline sodium citrate （SSC） as a spotting solution. Using a CY5-labeled DNA probe prepared through PCR amplification, the optimized glass slide hybridization could detect as little as 1.71 pg of tobacco mosaic virus （TMV） RNA. The sensitivity of the modified method was four times that of dot-blot hybridization on nylon membrane with a32P-labeled probe. The absence of false positive within the genus Potyvirus [potato virus A, potato virus Y （PVY） and zucchini yellow mosaic virus] showed that this method was highly specific. Furthermore, potato spindle tuber viroid （PSTVd） was also detected specifically. A test of 40 field potato samples showed that this method was equivalent to the conventional dot-blot hybridization for detecting PVY and PSTVd. To our knowledge, this is the first report of using dot-blot hybridization on glass slides with fluorescent-labeled probes for detecting plant RNA viruses and a viroid.

关键词： glass slide hybridization fluorescent labeling dot-blot hybridization virus viroid

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Quantitative Determination of Cucumber Mosaic Virus Genome RNAs in Virions by Real-Time Reverse Transcription-Polymerase Chain Reaction

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Acta Biochimica et Biophysica Sinica 2006年第10期38卷 669-676页

作者： Jun-Li FENG~1 Shao-Ning CHEN~1 Xiang-Shan TANG~1 Xian-Feng DING~1 Zhi-You DU~2 Ji-Shuang CHEN~(1,2*) 1 institute of bioengineering,zhejiang sci-tech university,hangzhou 310018,china 2 College of Life sciences,zhejiang university,hangzhou 310029,china institute of bioengineering Zhejiang Sci-Tech UniversityHangzhou 310018 China College of Life sciences Zhejiang UniversityHangzhou 310029 China

A real-time RT-PCR procedure using the green fluorescent dye SYBR Green Ⅰ was developedfor determining the absolute and relative copies of cucumber mosaic virus （CMV） genomic RNAs containedin purified *** specific to each CMV ORF were designed and *** were thenamplified with length varying from 61 to 153 *** dilution series of CMV genome RNAs prepared by invitro transcription as the standard samples,a good linear correlation was observed between their thresholdcycle （C_t） values and the logarithms of the initial template *** copies of genomic RNA 1,RNA 2,RNA 3 and the subgenomic RNA 4 in CMV virions were quantified by this method,and the ratios were about1.00:1.17:3.58:*** results were confirmed by Lab-on-a-Chip and Northern blot hybridization *** work is the first report concerning the relative amounts of different RNA fragments in CMV virions asa virus with tripartite genome.

关键词： real-time reverse transcription-polymerase chain reaction cucumber mosaic virus genomic RNA quantitative determination

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Satellite RNA-mediated Reduction of Cucumber Mosaic Virus Genomic RNAs Accumulation in Nicotiana tabacum

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Acta Biochimica et Biophysica Sinica 2007年第3期39卷 217-223页

作者： Qiansheng LIAO~(1,2) Liping ZHU~2 Zhiyou DU~2 Rong ZENG~(2,3) Junli FENG~2 Jishuang CHEN~(1,2*) 1 College of Life sciences,zhejiang university,Hangzhon 310029,china 2 institute of bioengineering,zhejiang sci-tech university,hangzhou 310018,china 3 College of Biological Safety technology Hunan Agricultural university,Changsha 410128,china College of Life sciences Zhejiang UniversityHangzhou 310029 China institute of bioengineering Zhejiang Sci-Tech UniversityHangzhou 310018 China College of Biological Safety technology Hunan Agricultural UniversityChangsha 410128 China

Satellite RNAs(satRNAs)are molecular parasites that interfere with the pathogenesis of thehelper *** this study,the relative accumulation of cucumber mosaic virus(CMV)-Fny genomicRNAs with or without satRNAs were quantitatively analyzed by real-time *** results showed thatsatRs apparently attenuated the symptoms of CMV-Fny on Nicotiana tabacum by depressing the accumu-lation of CMV-Fny genomic RNAs,tested as open reading *** accumulation of CMV-Fny 1a,2a,2b,3a,and CP genes was much higher than that of CMV-Fny with satRs added(CMV-Fsat),at differentinoculation ***-FnyΔ2b,in which the complete 2b gene and 41 amino acids at the C-terminal of the2a gene were deleted,caused only a slight mosaic effect on *** seedlings,similar to that of CMV-Fsat,but the addition of satRs to CMV-FnyΔ2b showed further decrease in the accumulation of CMV-FnyΔ2b genomic *** results indicated that the attenuation of CMV,by adding satRs or deleting the2b gene,was due to the low accumulation of CMV genomic RNAs,and that satRNA-mediated reduction ofCMV genomic RNAs accumulation in *** was possibly related to the 2b gene.

关键词： cucumber mosaic virus satellite RNA deletion of 2b gene relative accumulation

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Noether-type theory for discrete mechanico-electrical dynamical systems with nonregular lattices

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science china(Physics,Mechanics & Astronomy) 2010年第9期53卷 1687-1698页

作者： FU JingLi1,CHEN LiQun2 & CHEN BenYong3 1 institute of Mathematical Physics,zhejiang sci-tech university,hangzhou 310018,china 2 Department of Mechanics,Shanghai university,Shanghai 200072,china 3 Faculty of Mechanical-Engineering & Automation,zhejiang sci-tech university,hangzhou 310018,china institute of Mathematical Physics Zhejiang Sci-Tech University Hangzhou China Department of Mechanics Shanghai University Shanghai China Faculty of Mechanical-Engineering & Automation Zhejiang Sci-Tech University Hangzhou China

We investigate Noether symmetries and conservation laws of the discrete mechanico-electrical systems with nonregular *** operators of discrete transformation and discrete differentiation to the right and left are introduced for the *** on the invariance of discrete Hamilton action on nonregular lattices of the systems with the dissipation forces under the infinitesimal transformations with respect to the time,generalized coordinates and generalized charge quantities,we work out the discrete analog of the generalized variational *** this formula we derive the discrete analog of generalized Noether-type identity,and then we present the generalized quasi-extremal equations and properties of these equations for the *** also obtain the discrete analog of Noether-type conserved laws and the discrete analog of generalized Noether theorems for the *** we use an example to illustrate these results.

关键词： Noether symmetry variational formula quasi-extremal equation conservation law discrete mechanico-electrical dynamical system

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Recombinant Functional Human Lactoferrin Expressed in Baculovirus System

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Acta Biochimica et Biophysica Sinica 2006年第3期38卷 201-206页

作者： Tao LIU~(1,2) Yao-Zhou ZHANG~2* Xiang-Fu WU~31 institute of Biochemistry,College of Life sciences,zhejiang university,hangzhou 310029,china 2 institute of Biochemistry,zhejiang sci-Tec university,hangzhou 310018,china 3 institute of Biochemistry and Cell Biology,Chinese Academy of sciences Shanghai 200031,china institute of Biochemistry College of Life Sciences Zhejiang UniversityHangzhou 310029 China institute of Biochemistry Zhejiang Sci-Tec UniversityHangzhou 310018 China institute of Biochemistry and Cell Biology Chinese Academy of SciencesShanghai 200031 China

Human lactoferrin (hLf) is a multifunctional iron-binding *** this study,weamplified hLfcDNA by reverse transcription-polymerase chain reaction from normal human mammary *** nucleotide sequence of the hLf was identical to the known *** constructed a recombinant virus,vBm-hLf,harboring the hLf gene and exploited the BmN cells as host to produce recombinant human lactoferrin(rhLf).It was found that a recombinant protein with a molecular mass of approximately 78 kDa was *** 13.5μg rhLf was purified from 1-2×10～5 BmN cells infected by vBm-hLf and the rhLfproved to be biologically *** method established in our study will pave the way for efficient productionof rhLf for further application of this protein in the future.

关键词： bacteriostatic activity baculovirus Bombyx mori human lactoferrin reverse transcriptionpolymerase chain reaction

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Mutational Analysis of the Absolutely Conserved B8Gly: Consequence on Foldability and Activity of Insulin

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Acta Biochimica et Biophysica Sinica 2005年第10期37卷 673-679页

作者： Zhan-Yun GUO~1 Zhou ZHANG~1 Xiao-Yuan JIA~1 Yue-Hua TANG~(1,2) You-Min FENG~(1,2*) 1 Key Laboratory of Proteontics,institute of Biochemistry,and Cell Biology,Shanghai institutes for Biological sciences.Chinese Academy of sciences,Shanghai 200031.china 2 Instutute of Biochemistry,zhejiang sci-tech university,hangzhou 310018,china Key Laboratory of Proteomics Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of SciencesShanghai 200031 China institute of Biochemistry Zhejiang Sci-Tech UniversityHangzhou 310018 China

B8Gly is absolutely conserved in insulins during ***,its correspondingposition is always occupied by a Gly residue in other members of insulin *** work showedthat Ala replacement of BSGly significantly decreased both the activity and the foldability of *** effects of substitution are complicated,and different replacements sometimes cause significantly *** analyze the effects of B8 replacement by different amino acids,three new insulin/single-chaininsulin mutants with BSGly replaced by Ser,Thr or Leu were prepared by protein engineering,and both theirfoldability and activity were *** general,replacement of BSGly by other amino acids causes significantdetriment to the foldability of single-chain insulin:the conformations of the three B8 mutants are essentiallydifferent from that of wild-type molecules as revealed by circular dichroism;their disulfide stabilities inredox buffer are significantly decreased:their in vitro refolding efficiencies are decreased approximatelytwo folds;the structural stabilities of the mutants with Ser or Thr substitution are decreased significantly,while Leu substitution has little effect as measured by equilibrium guanidine *** far as biologicalactivity is *** replacement of B8Gly has only a moderate effect:its insulin receptor-bindingactivity is 23% of native *** Thr or Leu replacement produces significant detriment:the receptor-binding potencies of the two mutants are less than 0.2% of native *** present results suggest thatGly is likely the only applicable natural amino acid for the B8 position of insulin where both foldability andactivity are concerned.

关键词： insulin foldability activity disulfide stability

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Top 10 Influential Events in carbon neutrality and climate change response for 2023

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Frontiers in Energy 2024年第1期18卷 1-3页

作者： Research institute of Carbon Neutrality of Shanghai Jiao Tong university Research institute of Carbon Neutrality Shanghai Jiao Tong UniversityShanghai 200240China

In 2023, the global community intensified its efforts towards achieving carbon neutrality and addressing climate change, witnessing several impactful events. Research institute of Carbon Neutrality, Shanghai Jiao Tong university compiled the Top 10 Events (in chronological order) in the field of global carbon neutrality and climate change response for 2023, to share with readers these significant moments that document the ongoing ambition and collective efforts of human society.

关键词： neutral collective intensified

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Symmetries and exact solutions of discrete nonconservative systems

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science china(Physics,Mechanics & Astronomy) 2010年第9期53卷 1699-1706页

作者： FU JingLi1,LI XiaoWei2,LI ChaoRong1,ZHAO WeiJia3 & CHEN BenYong4 1 institute of Mathematical Physics,zhejiang sci-tech university,hangzhou 310018,china 2 Department of Physics,Shangqiu Normal university,Shangqiu 476000,china 3 Department of Mathematics,Qingdao university,Qingdao 266071,china 4 Faculty of Mechanical Engineering & Automation,zhejiang sci-tech university,hangzhou 310018,china institute of Mathematical Physics Zhejiang Sci-Tech University Hangzhou China Department of Physics Shangqiu Normal University Shangqiu China Department of Mathematics Qingdao University Qingdao China Faculty of Mechanical Engineering & Automation Zhejiang Sci-Tech University Hangzhou China

Based on the property of the discrete model entirely inheriting the symmetry of the continuous system,we present a method to construct exact solutions with continuous groups of transformations in discrete nonconservative *** Noether's identity of the discrete nonconservative system is *** symmetric discrete Lagrangian and symmetric discrete nonconservative forces are defined for the *** quasi-extremal equations of discrete nonconservative systems are *** conserved quantities are derived with symmetries associated with the continuous *** have also found that the existence of the one-parameter symmetry group provides a reduction to a conserved quantity;but the existence of a two-parameter symmetry group makes it possible to obtain an exact solution for a discrete nonconservative *** examples are discussed to illustrate these results.

关键词： discrete nonconservative system symmetry conserved quantity quasi-extremal equation exact solution

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Quasi-static Analysis of Thrust-loaded Angular Contact Ball Bearings Part Ⅱ:Results and Discussion

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Chinese Journal of Mechanical Engineering 2012年第1期25卷 81-87页

作者： CHEN Wenhua1,2,MA Zikui1,GAO Liang1,LI Xinglin3,and PAN Jun2 1 institute of Mechanical Design,zhejiang university,hangzhou 310027,china 2 institute of Mechanical Design and Manufacturing,zhejiang sci-tech university,hangzhou 310018,china 3 State Testing Laboratory of hangzhou Bearing Test & Research Center,hangzhou 310022,china institute of Mechanical Design Zhejiang University Hangzhou China institute of Mechanical Design and Manufacturing Zhejiang Sci-Tech University Hangzhou China State Testing Laboratory of hangzhou Bearing Test & Research Center Hangzhou China

Ball bearings are widely employed mechanical components characterized by high precision and quality,and usually play important roles in various rotary machines and *** advanced applications require a deep understanding of their various kinematic and tribological characteristics that are essential to predict the fatigue endurance,relieve the vibration and minimize the power dissipation of ball bearings in particular *** angular contact ball bearing under a specified operating condition is simulated with the quasi-static/creepage analytical model proposed in the preceding *** results demonstrate that the ball bearing is a statically determinate *** the balls spin on both inner and outer races means the ball is controlled by neither the inner nor the outer *** friction between the ball and raceway renders the inner and outer contact angles *** larger the coefficient of friction is,the larger the angle *** tangential traction perpendicular to the rolling direction due to the spin induces a gyro-like rotation of the ball with respect to the raceway even if no inertial effects are *** tangential elastic compliance of contacting surfaces gives rise to locked areas within the contact patch and transforms the sliding lines from circles into *** differential slip due to the close conformity of the ball and raceway makes the sliding and traction distributions asymmetric,which will influence the location of the spinning center of the ball with respect to the *** quasi-static/creepage model can be used to reveal the operating behaviors of ball bearings running under steady conditions and to optimize the design of ball bearings for specific applications.

关键词： ball bearing quasi-static creepage differential slip tangential elastic compliance

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Purification and Characterization of Two Endo-β-1,4-glucanases from Mollusca,Ampullaria crossean

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Acta Biochimica et Biophysica Sinica 2005年第10期37卷 702-708页

作者： Yan-Hong LI~(1#) Rui GUO~(3#) Qiu-Yu YIN~1 Ming DING~1 Si-Liang ZHANG~3 Gen-Jun XU~(1,2) Fu-Kun ZHAO~(1,2*) 1 Key Laboratory of Proteomics,institute of Biochemistry and Cell Biology,Shanghai institutes for Biological sciences.Graduate School of the Chinese Academy of sciences,Chinese Academy of sciences,Shanghai 200031,china 2 College of Life science,zhejiang sci-tech university,hangzhou 310018,china 3 bioengineering institute,East china university of science and technology,Shanghai 200233,china Key Laboratory of Proteomics Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Graduate School of the Chinese Academy of Sciences Chinese Academy of SciencesShanghai 200031 China bioengineering institute East China University of Science and TechnologyShanghai 200233 China College of Life science Zhejiang Sci-Tech UniversityHangzhou 310018 China

Two novel endo-β-1,4-glucanases,EG45 and EG27,were isolated from the gastric juice ofmollusca,Ampullaria crossean,by anion exchange,hydrophobic interaction,gel filtration and a secondround of anion exchange *** purified proteins EG45 and EG27 appeared as a single bandon sodium dodecylsulfate polyacrylamide gel electrophoresis with a molecular mass of 45 kDa and 27 kDa,*** optimum pH for CMC activity was 5.5 for EG45 and 4.4-4.8 for *** optimumtemperature range for EG27 was broad,between 50℃ and 60℃;for EG45 it was 50℃.The analysis on thestability of these two endo-β-1,4-glucanases showed that EG27 was acceptably stable at pH 3.0-11.0 evenwhen the incubation time was prolonged to 24 h at 30℃,whereas EG45 remained relatively stable at *** 85% of the activity of EG27 could be retained upon incubation at 60℃ for 24 ***,less than 10% residual activity of EG45 was detected at 50℃.Among different kinds of substrates,bothenzymes showed a high preference for carboxymethyl ***45,in particular,showed a carboxymethylcellulose hydrolytic activity of 146.5 IU/mg *** enzymes showed low activities to xylan (from oatspelt) and Sigmacell 101,and they were inactive to p-nitrophenyl-β-D-cellobioside,salicin and starch.

关键词： cellulase endo-β-1,4-glucanase Ampullaria crossean purification substrate specificity

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