多异瓢虫气味结合蛋白HvarOBP2的基因克隆及配体结合特性分析

【目的】本研究旨在克隆多异瓢虫Hippodamia variegata气味结合蛋白(odorant-binding proteins, OBPs)基因HvarOBP2,并解析这一蛋白的配体结合特征。【方法】采用BLAST和生物信息学技术从多异瓢虫成虫触角转录组中鉴定候选气味结合蛋白基因HvarOBP2,PCR特异扩增HvarOBP2基因全长cDNA序列;通过体外原核表达和亲和柱层析等手段获得HvarOBP2重组蛋白;运用荧光竞争结合实验测定重组蛋白HvarOBP2与40种植物挥发物组分以及24种蚜虫相关挥发物组分的结合能力;使用PyMOL1.9.0进行重组蛋白HvarOBP2与4种配体(β-紫罗兰酮、邻苯二甲酸二丁酯、油酸和橙花叔醇)的分子对接模拟。【结果】HvarOBP2(GenBank登录号：OK340816)开放阅读框(open reading frame, ORF)长447 bp,其编码蛋白N端有17个氨基酸组成的信号肽,具有6个保守的半胱氨酸位点,属于classical OBPs亚家族。荧光竞争结合实验结果表明,重组蛋白HvarOBP2与植物挥发物组分油酸(解离常数K     

A novel gene delivery system targeting cells expressing VEGF receptors

Two ligand oligopeptides GV1 and GV2 were designed according to the putative binding region of VEGF to its receptors.GV1,GV2 and endosome releasing oligopeptide HA20 were conjugated with poly-L-lysine or protamine and the resulting conjugates could interact with DNA in a noncovalent bond to form a complex.Using pSV2-β-galactosidase as a reporter gene,it has been demonstrated that exogenous gene was transferred into bovine aortic arch-derived endothelial cells (ABAE) and human malignant melanoma cell lines (A375) in vitro.In vivo experiments,exogenous gene was transferred into tumor vascular endothelial cells and tumor cells of subcutaneously transplanted human colon cancer LOVO,human malignant melanoma A375 and human hepatoma graft in nude mice.This system could also target gene to intrahepatically transplanted human hepatoma injected via portal vein in nude mice.These results are correlated with the relevant receptors(flt-1,flk-1/KDR) expression on the targeted cells and tissues.     

转基因小鼠用作脊髓灰质炎活疫苗神经毒力实验动物模型的研究

选用Ⅲ型脊髓灰质炎病毒,３批活疫苗样吕（ＷＨＯ／Ⅲ参考制品,９３／３６３和３Ｊ两批猴体神经毒力实验不合格的疫苗）和１株标准强毒株（Ｌｅｏｎ）,脊髓内注入携带有人细胞脊髓灰质炎病毒受体基因的转基因小鼠（ＰＶＲＴｇ２１）。临床和组织病理学检查表明,Ｌｅｏｎ病毒的毒力极强,２．０ｌｏｇ１０ＴＣＩＤ５０可使１００％小鼠麻痹和死亡,ＷＨＯ／Ⅲ疫苗参考制品毒力最弱,５．５ｌｏｇ１０ＣＩＤ５０才能使８１．７％小     

不同类型矮杨梅叶片中槲皮素含量比较

矮杨梅 (ＭｙｒｉｃａｎａｎａＣｈｅｖａｌ.)为杨梅科 (Ｍｙｒｉｃａｃｅａｅ)杨梅属 (ＭｙｒｉｃａＬ .)植物 ,自然分布于我国云南、贵州、西藏和四川的部分地区 ,为我国特有植物 ,具有药用价值和果品开发前景。近年来的研究表明 ,矮杨梅叶中含有黄酮类化合物槲皮素 (ｑｕｅｒｃｅｔｉｎ) ,有祛痰、止咳作用 ,还能对抗自由基 ,在抗癌、防癌、抗菌、消炎及心血管疾病的预防和治疗方面均具有较强的药理活性[1～ 4 ] 。虽然野生矮杨梅分布区范围窄 ,但其种内分化明显。刘宁等[5,6] 的研究表明 ,在矮杨梅根、茎、叶和果实等诸多器官的形态特征…     

Polyamine Changes and Chilling Injury in Cold-stored Loquat Fruits

Free polyamine levels (spermine (SPM), spermidine (SPD), and putrescine (PUT)) were determined using thin-layer chromatography and fluorometric method in loquat (Eriobotrya japonica Lindl. cv. Dahongpao) fruits stored at 1℃ and 12℃ and in postharvest SPM treated fruits stored at 1℃ respectively to investigate the relationship between changes in polyamines and chilling injury. In the loquat fruits stored at 1℃, SPM level decreased gradually in the first two weeks, then increased sharply and reached a peak value after three weeks, thereafter it decreased rapidly. SPD level decreased steadily during the first three weeks and increased significantly afterwards. PUT level evolved in a similar way as the SPM level did except that it increased slowly in the first two weeks. The fruit showed symptom of chilling injury manifested as flesh leatheriness after three weeks. However, no significant increase and decrease of these three polyamines was detected during storage at the nonchilling temperature (12℃). The SPM-treated fruits maintained high levels of SPM and SPD and remained low level of PUT during storage at 1℃, and no symptom of chilling injury was observed. These results suggested that the increase in SPM level in response to chilling exposure might serve as a defense mechanism against chilling injury while the accumulation of PUT could be a cause of the stress-induced injury and the increase in SPD level could be a consequence of this kind of stress.     

NaCl诱导大麦细胞微核及异常有丝分裂的研究

大麦幼苗生长在含NaCl的介质中[c(NaCl)=0, 0.05, 0.10, 0.15, 0.20, 0.25mol/L],其根尖细胞有丝分裂指数下降,各NaCl处理组的微核率均有不同程度地增加,并诱发产生了如染色体断片、后期染色体桥及"不均等分裂"等异常分裂。诱发微核及异常分裂细胞的比率与NaCl浓度和作用时间呈正相关,浓度提高或作用时间延长则微核千分率与异常分裂细胞的比率增高。     

重建端粒酶活性延长人成纤维细胞寿命的研究

正常人体细胞DNA的端粒随着细胞分裂而缩短,当缩短至一定长度时细胞将停止增殖并衰老死亡。细胞中的端粒酶对端粒起着补足长度的作用。但端粒酶在正常体细胞中不表达,只在生殖细胞、干细胞和肿瘤细胞中表达。最近已有将人端粒酶亚单位基因导入正常人体细胞而使细胞寿命延长的报道。本研究将人端粒酶催化亚基(hTERT)基因用电穿孔法转入正常人体成纤维细胞,筛选出阳性克隆后传代培养,确认外源性端粒酶基因表达和端粒酶活性的重建,证实细胞衰老延缓;同时,通过DNA整倍性和染色体核型分析,明确这些寿命延长的细胞并未发生恶性转化。目的在于通过在具有成骨潜能的成纤维细胞中重建端粒酶活性来延长它们作为骨修复种子细胞的寿命,并且对它们进一步用于临床的安全性进行考察。     

中国大萼苔科一新记录属

钝叶苔属(Ｃｌａｄｏｐｏｄｉｅｌｌａ)隶属于苔类植物的大萼苔科(Ｃｅｐｈａｌｏｚｉａｃｅａｅ),本属有2种:Ｃ.ｆｌｕｉｔａｎｓ(Ｎｅｅｓ)Ｓｐｒ.和Ｃ.ｆｒａｎｃｉｓｃｉ(Ｈｏｏｋ.)Ｂｕｃｈ.,广泛分布于欧洲和北美洲,其中前者在日本有记录。本属我国首次记录,仅发现1种:Ｃ.ｆｒａｎｃｉｓｃｉ(Ｈｏｏｋ.)Ｂｕｃｈ.,分布西藏。钝叶苔属ＣｌａｄｏｐｏｄｉｅｌｌａＢｕｃｈ.,Ｍｅｍ.Ｓｏｃ.Ｆ.Ｆｌ.Ｆｅｎｎｉｃａ5:87,1927.Ｔｙｐｅ:Ｃｌａｄｏｐｏｄｉｅｌｌａｆｌｕｉｔａｎｓ(Ｎｅｅｓ)Ｓｐｒ.植物体纤细,淡绿色。茎具腹面分枝,横切…     

Tet调控的EB病毒LMP1基因导入鼻咽癌细胞系表达的研究

本文利用Ｔｅｔ－ｏｎ基因表达系统建立了一株可诱导ＥＢ病毒ＬＭＰ１基因表达的鼻咽癌细胞株。首先将Ｔｅｔ－ｏｎ基因调控系统的调节质粒ｐＴｅｔ－ｏｎ导入鼻咽癌细胞系ＨＮＥ２中,用ｒｔＴＡ反应性芝光素酶报道基因ｐＴＲＥ－ｌｕｃ挑选高诱导、低背景的克隆,第二轮转染将构建的反应质粒ｐＴＲＥ－ＬＭＰ１导入得到稳定转染的阳性克隆,对各克隆用Ｗｅｓｔｅｒｎ印迹方法进行强力霉素诱导效应检测,其中Ｌ７对Ｄｏｘ具有良好的     

罗汉果叶中黄酮甙元的研究

以罗汉果叶为原料提取出黄酮,黄酮经水解、溶剂萃取、柱层析等手段,得到高纯度甙元;经TLC、液相色谱、红外光谱、核磁共振法等分析手段进行结构鉴定,证实甙元分别为山奈酚、槲皮素。