pEGFP-C1-GnRH/TRS在HeLa细胞中的遗传稳定性

目的研究质粒pEGFP-C1-GnRH/TRS在HeLa细胞中的遗传稳定性。方法取0代及传至第10、20、30和40代的工程细胞,在有选择压力(加G418)的条件下,测定pEGFP-C1-GnRH/TRS质粒丢失率。分别提取各代次工程细胞质粒DNA进行双酶切鉴定。将经酶切鉴定正确的第40代工程细胞质粒测序,并将各代次工程细胞加压筛选后,于荧光显微镜下观察。结果连续传10、20、30和40代后,质粒丢失率均不超过2%。不同代次的工程细胞质粒DNA经BamHⅠ/EcoRⅠ双酶切,酶切图谱相同。第40代质粒的GnRH/TRS序列与原代质粒相同。第40代细胞荧光分析与原代细胞相似。结论质粒pEGFP-C1-GnRH/TRS在HeLa细胞中有较好的遗传稳定性。     

Silicon analogues of the nonpeptidic GnRH antagonist AG-045572: syntheses, crystal structure analyses, and pharmacological characterization

AG‐045572 (CMPD1, 1 a ) is a nonpeptidic gonadotropin‐releasing hormone (GnRH) antagonist that has been investigated for the treatment of sex hormone‐related diseases. In the context of systematic studies on sila‐substituted drugs, the silicon analogue disila‐AG‐045572 ( 1 b ) and its derivative 2 were prepared in multi‐step syntheses and characterized by elemental analyses (C, H, N), NMR spectroscopic studies (¹H, ¹³C, ²⁹Si), and single‐crystal X‐ray diffraction. The pharmacological properties of compounds 1 a , 1 b , and 2 were compared in terms of their in vitro potency at cloned human and rat GnRH receptors. Compounds 1 a and 2 were also examined in regard to their pharmacokinetics and in vivo efficacy in both castrated rat (luteinizing hormone (LH) suppression) and intact rat (testosterone suppression) models. The efficacy and pharmacokinetic profiles of 1 a and its silicon‐containing analogue 2 appear similar, indicating that replacement of the 5,6,7,8‐tetrahydronaphthalene ring system by the 1,3‐disilaindane skeleton led to retention of efficacy. Therefore, the silicon compound 2 represents a novel drug prototype for the design of potent, orally available GnRH antagonists suitable for once‐daily dosing.     

人GnRH及其转运肽基因合成及鉴定

目的人工合成人GnRH及转运肽(TRS)基因。方法根据已发表的人GnRH基因mRNA序列以及转运肽(9个左旋精氨酸)基因核苷酸序列,设计一对核苷酸,再用含7mol/L尿素的变性聚丙烯酰胺凝胶电泳纯化,然后以这对核苷酸3′末端短互补序列退火、补齐,合成长达90bp的目的序列GnRH/TRS。将其亚克隆到pMD18T载体上,构建重组质粒pYC1。酶切鉴定筛选出阳性克隆pYC1,并测序。结果GnRH/TRS序列由90个核苷酸组成,扩增产物与原设计序列同源性达100%。结论已成功合成人GnRH/TRS基因,为进一步研究和应用奠定基础。     

基因工程菌株BLG8900的遗传稳定性

目的研究基因工程菌株BLG8900的遗传稳定性。方法在有选择压力(加氨苄西林)的条件下,测定pYC2质粒在大肠杆菌BLG8900株[含有质粒pYC2的BL21(DE3)]中的遗传稳定性。结果工程菌连续传10、25、50和100代后质粒具有良好的稳定性,而且经BamHⅠ/EcoRⅠ双酶切,酶切图谱相同。第100代菌株重组质粒的GnRH/TRS序列与原代菌株相同。原代与第10、25、50和100代菌株经IPTG诱导表达,GST-GnRH/TRS融合蛋白表达水平、菌体蛋白的SDS-PAGE图谱鉴定无明显差异。Western blot表明各代菌株表达产物均具有GnRH抗原特异性。结论质粒pYC2在大肠杆菌BL21(DE3)宿主菌中有较好的遗传稳定性。     

植物生产调节剂DCPTA的合成研究

在一步法合成DCPTA的基础上 ,通过加入相转移催化剂Bu4NBr ,使其收率提高到63.1% ,考察了最佳工艺条件。     

应用固定化金属亲和层析纯化His6融合甲状旁腺激素相关蛋白

应用原核细胞高效表达载体在大肠杆菌表达了N端带 6个组氨酸的重组人PTHrP。利用其对金属鳌合层析介质的亲合性 ,采用含盐咪唑梯度洗脱 ,一步纯化即可得到纯度大于 90 %且具有生物学活性的重组PTHrP ,并进一步制备了抗PTHrP单克隆抗体。应用His6纯化标签的固定化金属亲和层析 (IMAC) ,极大地方便了目的蛋白的纯化 ,为进一步研究PTHrP的生物学特性及其在临床诊断治疗中的应用奠定了基础。     

LHRH-PE40对黑色素瘤A375细胞的诱导凋亡作用

目的 研究促黄体激素释放激素-绿脓杆菌外毒素(Luteinizing hormone releasing hormone-Pseudomonas aeru-gionosa exotoxin 40,LHRH-PE40)对黑色素瘤A375细胞的诱导凋亡作用。方法用LHRH-PE40处理A375细胞,透射电镜观察细胞结构的改变;DNA ladder和流式细胞仪检测细胞的凋亡。结果经LHRH-PE40作用后,透射电镜观察可见A375细胞核聚集于一侧;DNA ladder检测发现,A375细胞的DNA电泳图像呈梯形条带;流式细胞术检测结果显示,A375细胞出现明显的二倍体峰,细胞周期也发生了相应的改变。结论 LHRH-PE40对A375细胞的细胞毒性作用可以有效诱导A375细胞凋亡。     

保幼激素及其类似物的研究与应用

归纳了保幼激素的合成与代谢研究,综述了保幼激素类似物的结构特点及其分类,并对保幼激素及其类似物在害虫防治、资源昆虫和其他方面的应用情况作了介绍.     

The so-called alkali-carbonate reaction (ACR) — Its mineralogical and geochemical details, with special reference to ASR

Typical examples of so-called alkali-carbonate reaction (ACR) in the Canadian field concretes in Ontario, CSA concrete prism, RILEM concrete microbars and RILEM mortar bar containing Pittsburg aggregate, were examined petrographically based on polarizing microscopy, SEM observation and quantitative SEM-EDS analysis of the reaction products. It was revealed that ASR gel was the main product responsible for the crack formation in concretes, and that this gel had a common nature to that in the typical ASR. That is, ASR gel presented distinctive compositional trend lines, passing from low-Ca ASR gel at [Ca/Si] = 1/2-1/6, [Ca]/[Na + K] = 1.0 to the “convergent point” with [Ca/Si] = 1.3-1.8, [Ca]/[Na + K] = 100 at which chemical equilibrium is attained with CSH gel. The so-called ACR is a combination of deleteriously expansive alkali-silica reaction (ASR) of cryptocrystalline quartz, and harmless dedolomitization which produces brucite and carbonate halo. In laboratory specimens, fine dolomitic aggregate undergoes dedolomitization, and brucite and ASR gel react to form non-expansive Mg-silicate gel on the dolomite crystals. This explains why the mortar bar produces smaller expansion than the concrete microbar, and why the reaction products are so minute that they escape attention by optical microscopy. As a crystalline counterpart, mountainite is a candidate for low-Ca ASR gel, while sepiolite is one for Mg-silicate gel. Concealed ASR was detected in ACR-affected field concretes undergoing ingress of deicing salt which formed Friedel's salt and Cl-doped CSH gel. Compositions of ASR products, methods of sample preparation and analysis for correct identification of ACR, and artifacts were critically reviewed.     

Synthesis, characterization, and properties of poly(ethylene terephthalate)/poly(1,4-butylene succinate) block copolymers

Reactive blending at 290 °C of a series of mixtures of poly(ethylene terephthalate) (PET) and poly(1,4-butylene succinate) (PBS) led to the formation of block PET/PBS copolyesters. The block lengths of the resulting copolymers decreased with the severity of the treatment. Copolyesters with PET/PBS molar compositions of 90/10, 80/20, 70/30, and 50/50 were prepared by this method and their composition and microstructure were characterized by ¹H and ¹³C NMR, respectively. The T_g, T_m, and crystallinity of the copolymers decreased as the content in PBS and the degree of randomness increased. The elastic modulus and tensile strength of the copolymers decreased with the content of PBS, whereas, on the contrary, the elongation at break increased. The PET/PBS copolymers exhibited a pronounced hydrolytic degradability, which increased with the content in 1,4-butylene succinic units. Hydrolysis mainly occurred on the aliphatic ester groups.     

In situ time-resolved X-ray diffraction of tobermorite formation in autoclaved aerated concrete: Influence of silica source reactivity and Al addition

The hydrothermal formation of tobermorite during the processing of autoclaved aerated concrete was investigated by in situ X-ray diffraction (XRD) analysis. High-energy X-rays from a synchrotron radiation source in combination with a newly developed autoclave cell and a photon-counting pixel array detector were used.To investigate the effects of the silica source, reactive quartz from chert and less-reactive quartz from quartz sand were used as starting materials. The effect of Al addition on tobermorite formation was also studied. In all cases, C-S-H, hydroxylellestadite and katoite were clearly observed as intermediates.Acceleration of tobermorite formation by Al addition was clearly observed. However, Al addition did not affect the dissolution rate of quartz. Two pathways, via C-S-H and katoite, were also observed in the Al-containing system. These results suggest that the structure of initially formed C-S-H is important for the subsequent tobermorite formation reactions.     

Crystallization behavior and hydrophilicity of poly(vinylidene fluoride)/poly(methyl methacrylate)/poly(vinyl pyrrolidone) ternary blends

Ternary blends composed of matrix polymer poly(vinylidene fluoride) (PVDF) with different proportions of poly(methyl methacrylate) (PMMA)/poly(vinyl pyrrolidone) (PVP) blends were prepared by melt mixing. The miscibility, crystallization behavior, mechanical properties and hydrophilicity of the ternary blends have been investigated. The high compatibility of PVDF/PMMA/PVP ternary blends is induced by strong interactions between the carbonyl groups of the PMMA/PVP blend and the CF₂ or CH₂ group of PVDF. According to the Fourier transform infrared and wide‐angle X‐ray difffraction analyses, the introduction of PMMA does not change the crystalline state (i.e. α phase) of PVDF. By contrast, the addition of PVP in the blends favors the transformation of the crystalline state of PVDF from non‐polar α to polar β phase. Moreover, the crystallinity of the PVDF/PMMA/PVP ternary blends also decreases compared with neat PVDF. Through mechanical analysis, the elongation at break of the blends significantly increases to more than six times that of neat PVDF. This confirms that the addition of the PMMA/PVP blend enhances the toughness of PVDF. Besides, the hydrophilicity of PVDF is remarkably improved by blending with PMMA/PVP; in particular when the content of PVP reaches 30 wt%, the water contact angle displays its lowest value which decreased from 91.4° to 51.0°. Copyright © 2011 Society of Chemical Industry