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Cloning of Ds MAPK Gene and Construction of Antisense Expression Vector

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Agricultural Biotechnology 2018年第4期7卷 8-11,24页

作者： jinrong yue Yuting CONG Xiangnan GAO Zhenyu XING Wenjing yue Xiaojie CHAI Dalian Ocean University/Key Laboratory of Maricuhure&Stock Enhancement in North China's Sea Ministry of Agricuhure/Key Laboratory of Marine Bio-Resources Restoration and Habitat Reparation in Liaoning ProvinceDalian 116023China

In order to investigate the role of MAPK gene in adaptation of Dunaliella salina to hypersaline environment, the Ds MAPK gene of D. salina was amplified by PCR. After inverted insertion of the open reading flame ofDs MAPK gene into downstream sequence of 35S promoter of plant expression vector, an antisense expression vector of Ds MAPK gene was successfully constructed and introduced into D. salina cells by LiAc/PEG-mediated method. The expression of Ds MAPK gene in transgenic D. salina was analyzed by semi-quantitative RT-PCR method. The results showed that the expression of Ds MAPK gene in D. salina was significantly inhibited at the transcriptional level. The study laid the foundation for further identification of the function of Ds MAPK gene.

关键词： Dunaliella salina Ds MAPK Antisense expression vector Semi-quantitative RT-PCR

Subcellular Localization of Small GTP-binding Protein DsRab in Dunaliella salina

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Agricultural Biotechnology 2018年第3期7卷 77-80页

作者： Yuting CONG jinrong yue Zhenyu XING Xiangnan GAO Xiujuan LI Xiaojie CHAI Dalian Ocean University Key Laboratory of Hydrobiology in Liaoning Province

With the total RNA of Dunaliella salina as a template,the cD NA sequence of D. salina small GTP-binding protein gene was amplified by RT-PCR technique,and cloned onto pM Dl8-T simple vector,the recon was subjected to PCR detection and restriction endonuclease analysis,and the total sequence of DNA was determined. The results showed that the cloned fragment was 612 bp,and shared 100% homology with reported D. salina DsRab gene（ GenB ank： JN989548）. The target gene fragment was inserted downstream of pM DCG 35 S promoter,constructing subcellular localization recombinant vector pM DCG-DsRab. The successfully constructed subcellular localization recombinant vector pM DCG-DsRab was transformed into Agrobacterium tumefaciens LBA4404,and positive single clones were screened and used for transinfection of onion epidemical cells by Agrobacterium-mediated method,and the instant expression of DsRab was observed under fluorescence microscope. The results showed that the fusion protein GFP-DsRab was successfully expressed in onion epidemical cells,and mainly distributed on cytomembrane. This study will provide reference for further illumination of the function and action mechanism of D. salina small GTP-binding protein DsRab.

关键词： Dunaliella salina Small GTP-binding proteins Subcellular localization Fusion protein

Cloning and Prokaryotic Expression of MAPKK kinase Gene from Dunaliella salina

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Cloning and Prokaryotic Expression of MAPKK kinase Gene from...

利用免疫共沉淀联合质谱技术筛选盐藻MAPK的互作蛋白

中国生物化学与分子生物学会2016年全国学术会议

作者： Chai Xiaojie Liu Yiqiong Cong Yuting Wang Yiyun yue jinrong Key Laboratory of Hydrobiology in Liaoning Province Dalian Ocean University

A MAPKK kinase gene was firstly cloned from Dunaliela salina by RT-PCR and RACE *** full length of c DNA for MAPKKK gene was 1 460 bp with a 27bp 5’UTR,a 554bp 3’UTR and a 879bp open reading frame coding 292 amino *... 详细信息

关键词： Dunaliella salina MAPKKK gene cloning bioinformatics prokaryotic expression western blotting

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核农学报 2020年第6期34卷 1187-1195页

作者：岳金荣丛玉婷邢震宇高相楠王明芳柴晓杰大连海洋大学/辽宁省省级高校水生生物学重点实验室辽宁大连116023

为进一步探究杜氏盐藻促有丝分裂原活化蛋白激酶(DsMAPK)的功能,采用免疫共沉淀联合质谱技术筛选盐藻MAPK的互作蛋白。将pGS21a-MAPK质粒转入大肠杆菌BL21中表达MAPK蛋白并制备多克隆抗体;将培养至对数生长期的盐藻细胞进行盐胁迫处理,... 详细信息

为进一步探究杜氏盐藻促有丝分裂原活化蛋白激酶(DsMAPK)的功能,采用免疫共沉淀联合质谱技术筛选盐藻MAPK的互作蛋白。将pGS21a-MAPK质粒转入大肠杆菌BL21中表达MAPK蛋白并制备多克隆抗体;将培养至对数生长期的盐藻细胞进行盐胁迫处理,然后提取盐藻总蛋白,并进行SDS-PAGE和Western blotting检测;以内源性靶蛋白为诱饵,将细胞总蛋白与MAPK抗体进行共孵育,将经蛋白A/G琼脂糖珠纯化的免疫共沉淀复合物进行质谱检测。结果表明,制备的多克隆抗体特异性良好;筛选出165种特有的差异蛋白。通过GO和KEGG分析发现,这些差异蛋白主要参与了新陈代谢、遗传信息的传递以及信号转导等生物学调控过程。蛋白质互作网络分析发现,直接与MAPK相互作用的蛋白有4种。本研究结果为深入研究杜氏盐藻响应盐胁迫的分子机制提供了参考。

关键词：杜氏盐藻 MAPK 免疫共沉淀技术质谱技术生物信息学

盐藻钙依赖蛋白激酶基因DsCDPK的表达分析

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水产科学 2019年第2期38卷 248-253页

作者：丛玉婷邢震宇岳金荣高相楠张晓琳柴晓杰大连海洋大学辽宁省省级高校水生生物学重点实验室辽宁大连116023

以盐藻总RNA为模板,利用RT-PCR技术扩增了盐藻DsCDPK基因的cDNA序列,将其克隆到pMD^(TM)19-T simple载体上,经测序获得的克隆片段全长1650 bp,与已发表的盐藻DsCDPK(GenBank:JQ964113)的编码区序列同源性达100%。将DsCDPK基因的开放阅... 详细信息

以盐藻总RNA为模板,利用RT-PCR技术扩增了盐藻DsCDPK基因的cDNA序列,将其克隆到pMD^(TM)19-T simple载体上,经测序获得的克隆片段全长1650 bp,与已发表的盐藻DsCDPK(GenBank:JQ964113)的编码区序列同源性达100%。将DsCDPK基因的开放阅读框与质粒pET-32a(+)连接,构建原核表达载体pET-32a-DsCDPK。将该重组质粒转化到大肠杆菌BL21中,经IPTG诱导融合,蛋白在大肠杆菌BL21中得到成功表达。通过SDS-PAGE检测发现,融合蛋白为部分可溶性表达,将上清蛋白经过His柱纯化后获得了纯度较高的可溶性融合蛋白,Western杂交检测显示,融合蛋白能被抗His单克隆抗体特异性识别,初步证明该融合蛋白就是带有His标签的DsCDPK蛋白。采用实时荧光定量PCR方法分析了高盐胁迫下DsCDPK基因的表达模式。试验结果表明,盐藻DsCDPK基因为盐胁迫上调基因,在高盐(3.0 mol/L NaCl)胁迫下,DsCDPK基因表达量显著增加,盐胁迫1 h时表达量达到最高,为正常生长状况(1.0 mol/L NaCl)下的3倍,差异达到极显著水平(P<0.01)。该研究成果为进一步阐明盐藻钙依赖蛋白激酶基因的功能及作用机制奠定了基础。

关键词：杜氏盐藻 DsCDPK 原核表达实时荧光定量PCR 盐胁迫

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南京东郊石炭纪威宁期的风暴沉积

资源调查与环境 1989年第1期21卷 75-84页

作者：邱金荣岳文浙业治铮地质矿产部南京地质矿产研究所

南京东郊石炭纪威宁期碳酸盐岩地层中普遍见有风暴沉积,以有突变的底界和底面构造、贝壳层、粒序层以及在风暴低能快速堆积条件下所形成的遮蔽构造等为基本鉴别要素,这些要素的不同组合构成的剖面结构可划分为五种类型,据此笔者提出本... 详细信息

南京东郊石炭纪威宁期碳酸盐岩地层中普遍见有风暴沉积,以有突变的底界和底面构造、贝壳层、粒序层以及在风暴低能快速堆积条件下所形成的遮蔽构造等为基本鉴别要素,这些要素的不同组合构成的剖面结构可划分为五种类型,据此笔者提出本区风暴沉积的综合剖面结构。

关键词：风暴沉积风暴岩南京东郊

Establishment of a New Method for Genetic Transformation of Dunaliella salina

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Agricultural Science & Technology 2017年第8期18卷 1374-1377页

作者：柴晓杰靳非丛玉婷岳金荣刘艺琼大连海洋大学/辽宁省水生生物学重点实验室辽宁大连116023

The exogenous gene was integrated into Dunaliella salina successfully by using LiAc/PEG mediating method for the first time. According to the results of histochemical staining, transgenic D. salina was blue, showing that the exogenous GUS gene was successfully expressed in the cells of D. salina. Meanwhile, the effects of growth state of D. salina, plasmid concentration and temperature on its transformation efficiency were studied, and the transformation conditions were optimized. The results show that the optimum conditions for the genetic transformation of D. salina are shown as follows： D. salina was in the early logarithmic phase; plasmid DNA concentration was 600 μg/ml; temperature was 29 ℃, and transformation efficiency was up to 74.8‰ under the best conditions. According to the results of PCR amplification and PCR-Southern hybridization, the target gene had been integrated into genome and was hereditary.

关键词： Dunaliela salina LiAc/PEG mediating method GUS gene Genetic transformation

Turning the V site in V@2D-BC_(3)N_(2)complex to high curvature state for efficient CO_(2)electroreduction to hydrocarbons

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Chinese Chemical Letters 2023年第8期34卷 328-332页

作者： Chaozheng He yue Yu Chenxu Zhao jinrong Huo Institute of Environmental and Energy Catalysis School of Materials Science and Chemical EngineeringXi’an Technological UniversityXi’an 710021China School of Sciences Xi’an Technological UniversityXi’an 710021China

Hydrocarbons are promising products for CO_(2)electroreduction(CRR)while is impeded by the low *** the curvature of the active site is an effective strategy to change the adsorption properties and further regulate the product distribution and ***,we have designed a novel V single atom catalyst(SAC)based on rolled two-dimensional(2D)BC_(3)N_(2)substrate with different *** results have demonstrated that increased curvature can enhance the adsorption strength of CRR intermediates,which follows different mechanisms for systems with low and high *** character eventually leads to the deviation away from the scaling line between Ead[CO]∼Ead[COOH]based on transition metals for V@2D-BC_(3)N_(2)systems.3-3 system is screened as the optimal candidate for hydrocarbons production due to the enhanced binding ability of adsorbates,which can increase the reactivity for hydrocarbons production and hinder the production of H2 and HCOOH simultaneously.

关键词： V doped 2D BC_(3)N_(2) Curvature effect CO_(2)electroreduction Hydrocarbons production d-band center theory Geometric and electronic properties

Antibacterial and antioxidant bifunctional hydrogel based on hyaluronic acid complex MoS_(2)-dithiothreitol nanozyme for treatment of infected wounds

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Regenerative Biomaterials 2024年第4期3卷 98-110页

作者： Yongping Lu Weiqi Kang yue Yu Ling Liang jinrong Li Haiying Lu Ping Shi Mingfang He yuemin Wang Jianshu Li Xingyu Chen Guangyuan Central Hospital Guangyuan 628000P.R.China State Key Laboratory of Polymer Materials Engineering College of Polymer Science and EngineeringSichuan UniversityChengdu 610065P.R.China Institute of Biomedical Engineering College of MedicineSouthwest Jiaotong UniversityChengdu 610031P.R.China

Wound repair is a complex physiological process that often leads to bacterial infections,which significantly threaten human ***,developing wound-healing materials that promote healing and prevent bacterial infections is *** this study,the coordination interaction between sulfhydryl groups on dithiothreitol(DTT)and MoS_(2)nanosheets is investigated to synthesize a MoS_(2)-DTT nanozyme with photothermal properties and an improved free-radical scavenging ***-bond-modified hyaluronic acid is used as a monomer and is cross-linked with a PF127-DA *** is prepared in coordination with MoS_(2)-DTT as the functional *** hydrogel exhibits antioxidant and antibacterial properties,attributed to the catalytic activity of catalase-like enzymes and photothermal *** the near-infrared(NIR),it exhibits potent antibacterial effects against gram-positive(Staphylococcus aureus)and gram-negative bacteria(Escherichia coli),achieving bactericidal rates of 99.76%and 99.42%,***,the hydrogel exhibits remarkable reactive oxygen species scavenging and antioxidant capabilities,effectively countering oxidative stress in L929 ***,in an animal model,wounds treated with the PHMoD(2.0)and NIR laser heal the fastest,sealing completely within 10 *** results indicate the unique biocompatibility and bifunctionality of the PHMoD,which make it a promising material for wound-healing applications.

关键词： hydrogel wound repair antibacterial antioxidant nanozyme photothermal therapy