一种基于Propidium Monoazide与PCR结合的选择性检测细菌活细胞的方法

建立了一种PMA与PCR结合的细菌活细胞检测的方法,可有效抑制微生物死细胞DNA的PCR扩增,从而排除死细胞对微生物活细胞检测的影响．结果表明,PMA浓度大于3 μg/ml时可抑制死细胞DNA的PCR扩增,PMA浓度高达50 μg/ml时仍不影响活细胞DNA的PCR扩增;并且得出曝光时间大于3 min可使PMA与死细胞DNA分子共价交联,抑制其PCR扩增;浊度影响PMA交联的结果表明浊度小于10 NTU时,PMA仍能有效地抑制死细胞DNA的PCR扩增,而当浊度大于100 NTU时,PMA失去效果．     

胜加端PCR技术改造hIGF—1cDNA

用ＤＮＡ合成仪合成了分别带有ＰｓＩ位点和ＳａｉＩ位点及张终止密码子的２个用于扩增ｈＩＤＧＦ１ｃＤＮＡ的ＰＣＲ引物，利用合成的引物，７００ｂｐ长的ｈＩＧＦ－Ｄ１ｃＤＮＡ模板和Ｔａｑ聚合酶进行ＰＣＲ扩增。     

水稻热激蛋白基因（HSP70）的PCR法快速分析

通过比较几个已知ＨＳＰ７０的ｃＤＮＡ顺序，用ＰＣＧＥＮＥ软件中的ＰＣＲＰＬＡＮ程序设计出一对简并引物，然后从水稻（广陆矮４号）总ＤＮＡ中扩增出ＨＳＰ７０的基因片段并将其克隆到ｐＢＬＵＥＳＣＲＩＰＴ载体中。在完成了对克隆的序列测定之后，用ＰＣＧＥＮＥ中有关程序对其进行同源分析，并对其他已知的ＨＳＰ７０作了一些分类及分子进化方面的探讨。     

输血后肝炎抗-HCV与HCV-RNA检测及临床意义

本文用套式聚合酶链反应及ELISA对66例输血后不同时期发生的输血后肝炎进行了HCV-RNA及抗-HCV的检测,43例抗-HCV阳性血清中,HCV-RNA阳性39例,23例抗-HCV阴性血清中,7例HCV-RNA阳性,按输血史时间长短分组测定结果表明,两项指标的消长情况不等,早期发病者中,以HCV-RNA阳性为主,晚期发病者中以抗-HCV为主。     

PCR—RFLP analysis of mitochondrial DNA ND5／6 region among3 subspecies of common carp（Cyprinus carpio L.）and its application to genetic discrimination of subspecies

Mitochondrial DNA ND5/6 region was studied by PCR-RFLP analysis among ten representative strains belonging to three subspecies (Cyprinus carpio carpio,Cyprinus carpio haematopterus and Cyprinus carpio rubrofuscus) of common carp(Cyprinus carpio L.)A total of 2.4kb fragment was amplified and subjected to restriction endonuclease analysis with nine restriction endonucleases subsequently.The results indicated that each subspecies owned one hyplotype and four restriction enzymes(Dde Ⅰ，HaeⅢ,Taq Ⅰ and MboⅠ)produced diagnostic restriction sites which could be used for discriminating the three subspecies and as molecular genetic markers for assistant selective breeding of common carp.     

多聚酶链反应技术在淋病检测上的应用

本文应用多聚酶链反应（ＰｏｌｇｍｅｒａｓｅＣｈａｉｎＲｅａｃｔｉｏｎ，ＰＣＲ）技术对直接涂片镜检淋球菌阴性患者１００例进行淋球菌ｃｐｐＢ基因片段测定。其中ＰＣＲ阳性者高达５４例，占５４％，该组病人中有２０例尖锐湿疣患者，其中ＰＣＲ阳性者１３例，占６５％。以上结果提示：ＰＣＲ技术可明显提高淋病患者的检出率。     

Overexpression of PITPNM3 promotes hepatocellular carcinoma cell metastasis

A previous study indicated that C–C chemokine(C–C motif)ligand 18(CCL18)is capable of inducing tumor cell invasion and metastasis by interacting with receptor membrane-associated phosphatidylinositol transfer protein 3(PITPNM3)in breast cancer cells.The present study aims to investigate the correlation between the PITPNM3 expression and metastasis in hepatocellular carcinoma(HCC).Real-time quantitative polymerase chain reaction and Western blot were performed to detect the expression pattern of PITPNM3 in patient samples and HCC cell lines.Wound-healing and transwell chamber assays were performed to assess the migration and invasiveness of HCC cells,and the activation of the signaling protein downstream of PITPNM3 was also detected by Western blot and immunofluorescence.The results revealed that PITPNM3 was upregulated in HCC tissue compared to matched normal liver tissue.Silencing the expression of PITPNM3 by specific siRNAs markedly attenuated the invasive and metastatic abilities of HCC cells,whereas the upregulation of PITPNM3 significantly increased HCC cell mobility.Furthermore,inhibiting the expression of PITPNM3 suppressed the activation of Pyk2,FAK,and Src,while overexpression of PITPNM3enhanced the phosphorylation of FAK and Src in HCC cells.Besides,suppression of Pyk2 can also impair the clustering of integrin.These results imply that PITPNM3 is a vital determinant of HCC migration and invasion.     

人胰岛素原类似物（B-Arg-Arg-A）基因的构建和表达

用聚合酶链反应（PCR）法将C肽为6个氨基酸残基的胰岛素原类似物基因删除突变成C肽仅为2个精氨酸残基的胰岛素原类似物（BArg-Arg-A）基因,即把pUC18BC'A改建为pUC18BR₂A。再将pUC18BR₂A与pJG105重组为表达质粒pJG107,使B-Arg-Arg-A与pJG105编码的一种多肽构成融合蛋白在大肠杆菌体系中表达。融合蛋白占细菌蛋白总量的58%。表达产物具有人胰岛素放射免疫活性。     

聚合酶链反应检测结核杆菌DNA的质量控制

根据结核杆菌（ＴＢ）ＤＮＡ的提取,耐热性多聚酶（Ｔａｑ）活性、结核Ｌ－型细菌的形成、变异、污染等,阐述了ＴＢ－ＤＮＡ多聚酶链式反应（ＰｏｌｙｍｅｒａｓｅＣｈａｉｎＲｅａｃ－ｔｉｏｎ,ＰＣＲ）检测中假阴性、假阳性出现的原因及内在规律,并提出了相应的改进措施。