The developmental stage-specific silencing of the human ε-globin gene during embryonic life is controlled, inpart, by the silencer (-392bp～-177bp) upstream of thisgene. In order to elucidate its role, the nuclear ex...The developmental stage-specific silencing of the human ε-globin gene during embryonic life is controlled, inpart, by the silencer (-392bp～-177bp) upstream of thisgene. In order to elucidate its role, the nuclear extractfrom the human fetal liver has been prepared and the interactions between trans-acting factors and this silencerelement have been examined. By using DNasel footprinting assay, a major protected region from -278bp to -235bpwithin this silencer element was identified. Furthermore,we found in gel mobility shift assay and Southwestern blotting assay that there were at least four trans-acting factors (MW ≈ 32, 28, 26 and 22kD) in the nuclear extractisolated from the human fetal liver, which could specifically bind to this region. Our results suggested that thesetrans-acting factors might play an important role in silencing the human embryonic ε-globin gene expression at thefetal stage through the interactions with this silencer.展开更多
Platymonas (Tetraselmis) subcordiformis is a unicellular marine green alga. It was found to be very sensitive to the herbicide Basta through a sensitivity test indicating it could be employed as a selective agent. The...Platymonas (Tetraselmis) subcordiformis is a unicellular marine green alga. It was found to be very sensitive to the herbicide Basta through a sensitivity test indicating it could be employed as a selective agent. The bar gene is a practicable and selectable marker gene. The vector containing the expression cassette of the bar gene was transferred to P. subcordiformis by both particle bombardment and glass-bead agitation and transformants were then selected using Basta. Finally, Southern blotting analysis indicated that the bar gene had been successfully integrated into the nuclear genome of P. subcordiformis using both of the transgenic techniques, with the transformation efficiency of the glass-bead method being slightly higher than that of particle bombardment. This is the first report on stable transformation of P. subcordiformis, and will improve fundamental research and enlarge application of this alga.展开更多
OBJECTIVE: To further characterize the differentiation inducing properties of EDRF1 and demonstrate its functional pathway involved in regulation of globin gene expression. METHODS: By transfecting EDRF1 sense and ant...OBJECTIVE: To further characterize the differentiation inducing properties of EDRF1 and demonstrate its functional pathway involved in regulation of globin gene expression. METHODS: By transfecting EDRF1 sense and antisense constructs into HEL cells, we identified the expression of globin and erythropoietin receptor genes by Northern blot analysis. RT-PCR and EMSA (electrophoresis mobility shift assay) were performed to monitor the expression and DNA-binding activity of erythroid specific transcription factors GATA-1 and NF-E2. RESULTS: It was shown that when EDRF1 was overexpressed, production of alpha-globin increased. In antisense EDRF1, overexpression of HEL cells, significant loss of alpha-, gamma-globin mRNA synthesis was observed. The transcription of endogenous GATA-1 and NF-E2 mRNA expression were maintained at the same levels compared with control experiments. However, the transcription activity of GATA-1 was severely impaired. Expression of erythropoietin receptor gene was not influenced by EDRF1 gene overexpression. CONCLUSION: The results suggested that EDRF1 regulated alpha- and gamma-globin gene synthesis by modulating DNA-binding activity of GATA-1 transcription factor.展开更多
MicroRNAs （miRNAs） are small non-coding RNAs that regulate gene expression at the post-transcriptional level, and their aberrant expression occurs during the development of malignant diseases. Recently, miRNAs have ...MicroRNAs （miRNAs） are small non-coding RNAs that regulate gene expression at the post-transcriptional level, and their aberrant expression occurs during the development of malignant diseases. Recently, miRNAs have been proposed as potential prognostic and predictive biomarkers for early diagnosis. However, a major obstacle in rapid miRNA analysis from real samples is the lack of ultrasensitive and quantitative techniques. In this regard, the use of chemiluminescence （CL） system offers a highly sensitive strategy for detecting miRNAs. In this article, an ultrasensitive approach has been established for the quantification ofmiRNAs, using magnetic beads （MBs） and alkaline phosphatase （AP）-based CL system. This technique depends on sandwich hybridization among MBs-labeled capture probes, target miRNAs and biotin-labeled reporter probes, conjugation of streptavidin-alkaline phosphatase （SA-AP） to biotin-labeled reporter probes, and CL detection of AP-linked targets. Detection of miR-21 with this technique demonstrated a high selectivity and an ultralow limit of detection （LOD） of 60 fM with an extraordinarily wide range of six orders of magnitudes. The quantitation could be achieved by direct detecting target miRNA in serum samples within a total time of 1.5 h and did not require reverse transcription and polymerase chain reaction （PCR） amplification. Therefore, this developed method shows great potential for early cancer diagnosis based on miRNAs as biomarkers.展开更多
In order to provide scientific support to policy makers in the regulation of PM2.5 pollution in China, it is important to accurately assess the current status, spatiotemporal characteristics and regionalization data f...In order to provide scientific support to policy makers in the regulation of PM2.5 pollution in China, it is important to accurately assess the current status, spatiotemporal characteristics and regionalization data for this air pollutant. An analysis of the pollution status of PM2.5 was conducted using daily averaged mass concentration data recorded in 74 cities in 2013 and 161 cities in 2014. The rotated empirical orthogonal function(REOF) method was applied to analyze this data. Results showed that the average annual PM2.5 concentration in urban areas of China is 62.2±21.5 ?g/m3, and that the distribution is spatially heterogeneous. The North China Plain, middle and lower Yangtze River Plain, Sichuan Basin and Guanzhong Plain had relatively high annual PM2.5 concentrations compared with the southeast coastal region, the Tibetan Plateau and the Yungui Plateau. PM2.5 mass concentrations tended to be higher in winter than in summer, however, the data for many cities showed a small peak in concentrations from May to July. An analysis of the spatial correlation of PM2.5 indicated a significant influence of topographic conditions. A lower correlation was observed where terrain features varied greatly. Based on the results of the REOF analysis and topographic characteristics, ten regions were identified in mid-eastern China, which could be considered as basic pollution prevention divisions for PM2.5; these include the North China Plain region, Pearl River Delta region, Jianghuai Plain region, middle Yangtze River Plain region, Northeast Plain region, Jiangnan coastal region, Sichuan Basin region, Qiantao Plain region, Guanzhong-Central Plain region and Yungui Plateau region. Seasonal variations in the regionalization data were observed, especially for the North China Plain and Pearl River Delta regions. Among the ten regions identified in this study, the North China Plain, Guanzhong-Central Plain, middle Yangtze River Plain and Jianghuai Plain had relatively high PM2.5 mass concentrations in comparison with the others. Therefore, these regions should be considered as the key regions to target in developing PM2.5 pollution prevention strategies. This study improves the present understanding of the spatial distribution, seasonal changes and regional status of PM2.5 pollution in China and helps establish possible control strategies for the reduction of this air pollutant.展开更多
文摘The developmental stage-specific silencing of the human ε-globin gene during embryonic life is controlled, inpart, by the silencer (-392bp～-177bp) upstream of thisgene. In order to elucidate its role, the nuclear extractfrom the human fetal liver has been prepared and the interactions between trans-acting factors and this silencerelement have been examined. By using DNasel footprinting assay, a major protected region from -278bp to -235bpwithin this silencer element was identified. Furthermore,we found in gel mobility shift assay and Southwestern blotting assay that there were at least four trans-acting factors (MW ≈ 32, 28, 26 and 22kD) in the nuclear extractisolated from the human fetal liver, which could specifically bind to this region. Our results suggested that thesetrans-acting factors might play an important role in silencing the human embryonic ε-globin gene expression at thefetal stage through the interactions with this silencer.
基金Supported by the National Basic Research Program of China (973 Program) (No. 2011CB200902)the National High Technology Research and Development Program of China (863 Program) (Nos. 2009AA10Z106,2009AA064401)+2 种基金the Promotive Research Fund for Excellent Young and Middle-aged Scientists of Shandong Province (No. 2010BSB02009)the National Natural Science Foundation of China (Nos. 40876082,30870247)the Science and Technology Project of Qingdao City (No.09-1-3-59-jch)
文摘Platymonas (Tetraselmis) subcordiformis is a unicellular marine green alga. It was found to be very sensitive to the herbicide Basta through a sensitivity test indicating it could be employed as a selective agent. The bar gene is a practicable and selectable marker gene. The vector containing the expression cassette of the bar gene was transferred to P. subcordiformis by both particle bombardment and glass-bead agitation and transformants were then selected using Basta. Finally, Southern blotting analysis indicated that the bar gene had been successfully integrated into the nuclear genome of P. subcordiformis using both of the transgenic techniques, with the transformation efficiency of the glass-bead method being slightly higher than that of particle bombardment. This is the first report on stable transformation of P. subcordiformis, and will improve fundamental research and enlarge application of this alga.
文摘OBJECTIVE: To further characterize the differentiation inducing properties of EDRF1 and demonstrate its functional pathway involved in regulation of globin gene expression. METHODS: By transfecting EDRF1 sense and antisense constructs into HEL cells, we identified the expression of globin and erythropoietin receptor genes by Northern blot analysis. RT-PCR and EMSA (electrophoresis mobility shift assay) were performed to monitor the expression and DNA-binding activity of erythroid specific transcription factors GATA-1 and NF-E2. RESULTS: It was shown that when EDRF1 was overexpressed, production of alpha-globin increased. In antisense EDRF1, overexpression of HEL cells, significant loss of alpha-, gamma-globin mRNA synthesis was observed. The transcription of endogenous GATA-1 and NF-E2 mRNA expression were maintained at the same levels compared with control experiments. However, the transcription activity of GATA-1 was severely impaired. Expression of erythropoietin receptor gene was not influenced by EDRF1 gene overexpression. CONCLUSION: The results suggested that EDRF1 regulated alpha- and gamma-globin gene synthesis by modulating DNA-binding activity of GATA-1 transcription factor.
基金supported by the National Key Program for Developing Basic Research (2014CB744501)the National High Technology Research and Development Program of China (2012AA022703)+8 种基金the National Key Special Science Program (2013ZX10004103-002)the National Natural Science Foundation of China (61471168, 61527806, 61271056)the Special Projects in Jiangsu Province (BL2014094)the Economical Forest Cultivation and Utilization of 2011 Collaborative Innovation Center in Hunan Province [(2013) 448]the Talents Planning of Six Summit Fields of Jiangsu Province (2013-WSN-056)Tianjin Medical University General Hospital Funding (ZYYFY2015029)China Postdoctoral Science Foundation funded project (2015T80487)Natural Science Foundation of Jiangsu Province (BK20140900)the Open Project of State Key Laboratory of Bioelectronics (2014HX12)
文摘MicroRNAs （miRNAs） are small non-coding RNAs that regulate gene expression at the post-transcriptional level, and their aberrant expression occurs during the development of malignant diseases. Recently, miRNAs have been proposed as potential prognostic and predictive biomarkers for early diagnosis. However, a major obstacle in rapid miRNA analysis from real samples is the lack of ultrasensitive and quantitative techniques. In this regard, the use of chemiluminescence （CL） system offers a highly sensitive strategy for detecting miRNAs. In this article, an ultrasensitive approach has been established for the quantification ofmiRNAs, using magnetic beads （MBs） and alkaline phosphatase （AP）-based CL system. This technique depends on sandwich hybridization among MBs-labeled capture probes, target miRNAs and biotin-labeled reporter probes, conjugation of streptavidin-alkaline phosphatase （SA-AP） to biotin-labeled reporter probes, and CL detection of AP-linked targets. Detection of miR-21 with this technique demonstrated a high selectivity and an ultralow limit of detection （LOD） of 60 fM with an extraordinarily wide range of six orders of magnitudes. The quantitation could be achieved by direct detecting target miRNA in serum samples within a total time of 1.5 h and did not require reverse transcription and polymerase chain reaction （PCR） amplification. Therefore, this developed method shows great potential for early cancer diagnosis based on miRNAs as biomarkers.
基金supported by the Special Funds of the State Environmental Protection Public Welfare Industry(201509001,201309016)the Transformation Project of China National Environmental Monitoring Center(2011ZX-010-001)
文摘In order to provide scientific support to policy makers in the regulation of PM2.5 pollution in China, it is important to accurately assess the current status, spatiotemporal characteristics and regionalization data for this air pollutant. An analysis of the pollution status of PM2.5 was conducted using daily averaged mass concentration data recorded in 74 cities in 2013 and 161 cities in 2014. The rotated empirical orthogonal function(REOF) method was applied to analyze this data. Results showed that the average annual PM2.5 concentration in urban areas of China is 62.2±21.5 ?g/m3, and that the distribution is spatially heterogeneous. The North China Plain, middle and lower Yangtze River Plain, Sichuan Basin and Guanzhong Plain had relatively high annual PM2.5 concentrations compared with the southeast coastal region, the Tibetan Plateau and the Yungui Plateau. PM2.5 mass concentrations tended to be higher in winter than in summer, however, the data for many cities showed a small peak in concentrations from May to July. An analysis of the spatial correlation of PM2.5 indicated a significant influence of topographic conditions. A lower correlation was observed where terrain features varied greatly. Based on the results of the REOF analysis and topographic characteristics, ten regions were identified in mid-eastern China, which could be considered as basic pollution prevention divisions for PM2.5; these include the North China Plain region, Pearl River Delta region, Jianghuai Plain region, middle Yangtze River Plain region, Northeast Plain region, Jiangnan coastal region, Sichuan Basin region, Qiantao Plain region, Guanzhong-Central Plain region and Yungui Plateau region. Seasonal variations in the regionalization data were observed, especially for the North China Plain and Pearl River Delta regions. Among the ten regions identified in this study, the North China Plain, Guanzhong-Central Plain, middle Yangtze River Plain and Jianghuai Plain had relatively high PM2.5 mass concentrations in comparison with the others. Therefore, these regions should be considered as the key regions to target in developing PM2.5 pollution prevention strategies. This study improves the present understanding of the spatial distribution, seasonal changes and regional status of PM2.5 pollution in China and helps establish possible control strategies for the reduction of this air pollutant.
