Gut Microbiota-Produced Tryptamine Activates an Epithelial G-Protein-Coupled Receptor to Increase Colonic Secretion

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猪血清白蛋白基因5′-端调控区的克隆与分析

根据已报道的hsa(人血清白蛋白基因)启动子的序列设计了一对引物,以猪的基因组DNA为模板,采用PCR扩增出一条256 bp的DNA链,用Primer Premier5.0和Promoter ScanⅡ软件进行分析。结果表明,此序列为psa基因5′-端上游调控区,该区域含有一般启动子的CAAT-box、TATA-box等基本的顺式作用元件。另外,还含有CTF/NF-1,CTF,APF,HNF-1, CP1等转录因子的结合位点。     

Constitutive expression of human coagulating factor IX in HeLa cells by homologous recombination of the promoter

Constitutive expression of hFIX protein in nonhepatocytes was studied. The gene targeting vector was constructed and transferred into HeLa cells. With the detection system of PCR, we demonstrated that the endogenoushFIX promoter was replaced with anhCMV promoter when targeted insertion of the constructor was directed by the sequence homology. The expression ofhFIX in the modified HeLa cells, 11.2 ng/10⁶ cell/24 h, strongly suggested thathFIX gene could be activated by a powerful promoter in nonhepatocytes. The results would make it possible to examine the feasibility of re-regulate gene expression by promoter replacement.     

Differentiated Gene Expression in Cells within Yeast Colonies

Yeast cells growing on solid media organize themselves into multicellular structures, colonies, exhibiting patterns specific for particular yeast strains. With the aim of identifying genes involved in regulations of the colony formation, we applied a new approach enabling the extensive screening of Saccharomyces cerevisiae genes, the expression of which is changed during colony development. We used the library of S. cerevisiae DNA fragments inserted in front of the lacZ gene lacking its own promoter. Colonies of transformants with a blue/white patterned morphotype, implying that the expression of the lacZ gene from the inserted yeast promoter is switched on and off during the colony formation, were isolated. We identified several genes with variable expression during colony morphogenesis, including CCR4, PAM1, MEP3, ADE5,7 and CAT2. S. cerevisiae strain deleted in the CCR4 gene forms colonies with less organized morphology when compared with the isogenic parental strain. The synchronization of the expression patterns of some of the isolated genes in neighboring colonies was observed.     

Tissue-dependent enhancement of transgene expression by introns of replacement histone H3 genes of Arabidopsis

Intron-bearing replacement histone H3 genes in Arabidopsis and other plants are highly and constitutively expressed. We demonstrate that the introns located within the 5-untranslated regions (5-UTR) of the two Arabidopsis replacement H3 genes will abolish the cell cycle dependence of an endogenous histone H4 promoter. We demonstrate that these introns, functionally combined with their endogenous promoters, could produce the high and constitutive expression of the replacement H3 genes observed in planta. They strongly increase gene expression whatever the promoter, from the strong 35S CaMV promoter to complete and resected promoters of cell cycle-dependent and replacement histone genes. Quantitative analysis of the extent of reporter gene enhancement in different parts of developing transgenic plantlets, ranging from 2-fold to 70-fold, supports the notion that trans-acting factors are responsible for this effect. Such factors appear most abundant in roots.     

nifH—gfp表达载体的构建及其在Enterobacter gergoviae 57—7中的表达

采用ＰＣＲ技术,从ＧＦＰｍｕｔ２中扩增得到三位点突变的报告基因ｇｆｐＳ６５Ｔ、Ｖ６８Ｌ、Ｓ７２Ａ片段,并将它和肺炎克氏杆菌（Ｋｌｅｂｓｉｅｌｌａ ｐｎｅｕｍｏｎｉａｅ（Ｓｃｈｒｏｅｅｔｅｒ）Ｔｒｅｖｉｓａｎ）Ｍ５ａ１的固氮酶结构基因ｎｉｆＨ的启动子和其起始密码子相融合,获得ｎｉｆＨ－ｇｆｐ表达载体ｐＭＧＦＰ２;再在ｐＭＧＦＰ２上插入卡那霉素抗性基因,获得可在日勾维肠杆菌（Ｅｎｔｅｒｏｂａｃｔｅｒ     

Expression of Acidothermus cellulolyticus endoglucanase E1 in transgenic tobacco: biochemical characteristics and physiological effects

The expression of the Acidothermus cellulolyticus endoglucanase E1 gene in transgenic tobacco (Nicotiana tabacum) was examined in this study, where E1 coding sequence was transcribed under the control of a leaf specific Rubisco small subunit promoter (tomato RbcS-3C). Targeting the E1 protein to the chloroplast was established using a chloroplast transit peptide of Rubisco small subunit protein (tomato RbcS-2A) and confirmed by immunocytochemistry. The E1 produced in transgenic tobacco plants was found to be biologically active, and to accumulate in leaves at levels of up to 1.35% of total soluble protein. Optimum temperature and pH for E1 enzyme activity in leaf extracts were 81°C and 5.25, respectively. E1 activity remained constant on a gram fresh leaf weight basis, but dramatically increased on a total leaf soluble protein basis as leaves aged, or when leaf discs were dehydrated. E1 protein in old leaves, or after 5h dehydration, was partially degraded although E1 activity remained constant. Transgenic plants exhibited normal growth and developmental characteristics with photosynthetic rates similar to those of untransformed SR1 tobacco plants. Results from these biochemical and physiological analyses suggest that the chloroplast is a suitable cellular compartment for accumulation of the hydrolytic E1 enzyme.