目的：为了探讨Rap2c基因与肺癌发生的关系，克隆人Ras家族小G蛋白Rap2c的cDNA，构建其真核表达质粒并在人肺癌细胞株A549和H1299中表达。方法人骨肉瘤细胞株U2OS提取细胞总RNA，经逆转录聚合酶链反应（RT-PCR）逆转录成cDNA，PCR扩增Ra...目的：为了探讨Rap2c基因与肺癌发生的关系，克隆人Ras家族小G蛋白Rap2c的cDNA，构建其真核表达质粒并在人肺癌细胞株A549和H1299中表达。方法人骨肉瘤细胞株U2OS提取细胞总RNA，经逆转录聚合酶链反应（RT-PCR）逆转录成cDNA，PCR扩增Rap2c，酶切后插入pcDNA3．1（＋）构建真核表达质粒 pcD-NA3．1（＋）-Rap2c，采用酶切及测序鉴定。重组质粒转染A 549和H 1299细胞，Western blot 检测其目的基因表达。结果双酶切及测序结果显示重组质粒pcDNA3．1（＋）-Rap2c成功构建，Werstern blot 检测到 A549和H 1299细胞有相应蛋白表达。结论成功构建人Rap2c基因真核表达质粒，为后续研究奠定了基础。展开更多
[目的]探讨RAS癌基因家族成员RAP2B(member of RAS oncogene family,RAP2B)在食管癌中的表达与临床诊断意义。[方法]采用免疫组化S-P法对87例食管癌、34例Barrett食管和15例正常食管组织进行检测,观察RAP2B在各组织中的表达。分析RAP2B...[目的]探讨RAS癌基因家族成员RAP2B(member of RAS oncogene family,RAP2B)在食管癌中的表达与临床诊断意义。[方法]采用免疫组化S-P法对87例食管癌、34例Barrett食管和15例正常食管组织进行检测,观察RAP2B在各组织中的表达。分析RAP2B的表达与食管癌患者性别、年龄、临床分型、组织学类型、TNM分期、肿瘤大小、淋巴转移的关系。[结果]RAP2B基因在食管癌、Barrett食管及正常食管黏膜组织中的阳性表达率分别为66.7%(58/87)、58.8%(20/34)、26%(4/15),该基因在食管癌组织中表达阳性率明显高于Barrett食管和正常食管组织,差异有统计学意义(χ^(2)=8.592,P<0.05);食管癌组织中RAP2B蛋白的阳性表达与肿瘤分化程度(χ^(2)=7.578,P<0.05)、淋巴转移(χ^(2)=5.331,P<0.05)有关,而与性别、年龄、肿瘤大小、TNM分期均无关(P>0.05)。[结论]RAP2B基因在食管癌组织中的过表达可能与食管癌的发生有关,其表达与组织学分级相关。展开更多
Based upon analysis of data obtained from theATH1 microarrays, a cDNA that was highly induced afterdrought treatment, was isolated from Arabidopsis seedlings.RT-PCR and Quantitative Real-Time (QRT)-PCR experi-ments sh...Based upon analysis of data obtained from theATH1 microarrays, a cDNA that was highly induced afterdrought treatment, was isolated from Arabidopsis seedlings.RT-PCR and Quantitative Real-Time (QRT)-PCR experi-ments showed that expression level of the gene increasedsignificantly upon drought, UV, abscisic acid, high salinityand salicylic acid treatments. It was classified as a DREBsubfamily member based on multiple sequence alignmentand phylogenetic characterization. Since it encoded a proteinwith a typical ERF/AP2 DNA-binding domain and containeda glutamine-rich region near its N terminus, we named itQRAP2 (for glutamine-rich AP2). Gel retardation assay re-vealed that QRAP2 was able to form a specific complex withthe previously characterized DRE element while did notshow any affinity to the GCC box or the mutant DRE box.When fused to the GAL4 DNA-binding domain, either full-length QRAP2 or its N-terminus functioned effectively as atrans-activator in the yeast one-hybrid assay with its C-ter-minus completely inactive. Our data indicate that QRAP2could be a new member of the AP2/EREBP transcriptionfactor family involved in activation of down-stream targetgenes in response to environmental stress, especially underdrought conditions.展开更多
文摘目的：为了探讨Rap2c基因与肺癌发生的关系，克隆人Ras家族小G蛋白Rap2c的cDNA，构建其真核表达质粒并在人肺癌细胞株A549和H1299中表达。方法人骨肉瘤细胞株U2OS提取细胞总RNA，经逆转录聚合酶链反应（RT-PCR）逆转录成cDNA，PCR扩增Rap2c，酶切后插入pcDNA3．1（＋）构建真核表达质粒 pcD-NA3．1（＋）-Rap2c，采用酶切及测序鉴定。重组质粒转染A 549和H 1299细胞，Western blot 检测其目的基因表达。结果双酶切及测序结果显示重组质粒pcDNA3．1（＋）-Rap2c成功构建，Werstern blot 检测到 A549和H 1299细胞有相应蛋白表达。结论成功构建人Rap2c基因真核表达质粒，为后续研究奠定了基础。
文摘[目的]探讨RAS癌基因家族成员RAP2B(member of RAS oncogene family,RAP2B)在食管癌中的表达与临床诊断意义。[方法]采用免疫组化S-P法对87例食管癌、34例Barrett食管和15例正常食管组织进行检测,观察RAP2B在各组织中的表达。分析RAP2B的表达与食管癌患者性别、年龄、临床分型、组织学类型、TNM分期、肿瘤大小、淋巴转移的关系。[结果]RAP2B基因在食管癌、Barrett食管及正常食管黏膜组织中的阳性表达率分别为66.7%(58/87)、58.8%(20/34)、26%(4/15),该基因在食管癌组织中表达阳性率明显高于Barrett食管和正常食管组织,差异有统计学意义(χ^(2)=8.592,P<0.05);食管癌组织中RAP2B蛋白的阳性表达与肿瘤分化程度(χ^(2)=7.578,P<0.05)、淋巴转移(χ^(2)=5.331,P<0.05)有关,而与性别、年龄、肿瘤大小、TNM分期均无关(P>0.05)。[结论]RAP2B基因在食管癌组织中的过表达可能与食管癌的发生有关,其表达与组织学分级相关。
基金This work was supported by the National Natural Science Foundation of China(Grant No.30221 120261).
文摘Based upon analysis of data obtained from theATH1 microarrays, a cDNA that was highly induced afterdrought treatment, was isolated from Arabidopsis seedlings.RT-PCR and Quantitative Real-Time (QRT)-PCR experi-ments showed that expression level of the gene increasedsignificantly upon drought, UV, abscisic acid, high salinityand salicylic acid treatments. It was classified as a DREBsubfamily member based on multiple sequence alignmentand phylogenetic characterization. Since it encoded a proteinwith a typical ERF/AP2 DNA-binding domain and containeda glutamine-rich region near its N terminus, we named itQRAP2 (for glutamine-rich AP2). Gel retardation assay re-vealed that QRAP2 was able to form a specific complex withthe previously characterized DRE element while did notshow any affinity to the GCC box or the mutant DRE box.When fused to the GAL4 DNA-binding domain, either full-length QRAP2 or its N-terminus functioned effectively as atrans-activator in the yeast one-hybrid assay with its C-ter-minus completely inactive. Our data indicate that QRAP2could be a new member of the AP2/EREBP transcriptionfactor family involved in activation of down-stream targetgenes in response to environmental stress, especially underdrought conditions.
