Science Letters: Quantification of Helicobacter pylori levels in soil samples from public playgrounds in Spain

Helicobacter pylori are ubiquitous Gram-negative bacteria with a high estimated level of infection in the world populations, but a majority of the infected persons are asymptomatic. This pathogen has been classified by the World Health Organization as a class I carcinogen and recognized as the causal agent of most peptic ulcers and chronic gastritis that might lead to stomach cancer. Although not all the transmission pathways of these bacteria into humans have been properly identified, enough data have sugg...     

影响聚式酶链式反应实验效果的基本因素

研究聚式酶链式反应(PCR)实验中影响其扩增效果的多种因素,寻求反应体系中理想的各因素浓度配比。选取含5+10优质亚基的小麦品种为材料,以特异扩增1DX5基因PCR实验为例,从模板DNA、引物终浓度、Taq酶、二价镁离子、4dNTPs混合液、缓冲液浓度几方面入手,通过设计梯度实验,研究不同因素对PCR扩增效果的影响。结果表明,反应体系6个基本因素,缺少任何1个都扩增不出产物。在反应总体积25μL的PCR反应体系中,模板DNA选用32 ng/μL,Mg2+终浓度2 mmol/L,Taq酶1.2 U/25μL,引物0.8μmol/μL,4dNTPs0.3 mmol/L,缓冲液1×buffer最为合适,能扩增出理想的结果。     

聚合酶链反应检测真菌感染的实验研究

本研究以NS5-NS6为引物,扩增真菌同源性序列ssu-rDNA片段,建立广范围真菌检测的方法。用此方法扩增医学主要致病真菌、细菌和人体细胞,结果所有真菌均扩增出一个约310bp的产物,而细菌和人体细胞均扩增阴性,1pg白色假丝酵母菌DNA即可检出。     

计算机软件在PCR实验上的应用

对与PCR实验紧密相关的计算机软件——引物设计软件、凝胶图像处理软件、数据分析软件和文献查找软件的应用进行了概述。     

聚合酶链反应检测儿童急性呼吸道流感嗜血杆菌感染的研究

目的 :建立儿童急性呼吸道感染的PCR检测方法 ,并与常规方法进行比较。方法 :选择编码外膜蛋白P6基因作为靶片段设计引物 ,分别用PCR方法及常规培养法检测 78例急性呼吸道感染儿童。结果 :建立的PCR检测法可以检测 10～ 5 0个流感嗜血杆菌 ;78例急性呼吸道感染儿童中PCR方法检出 4 2株 (5 3.8% ) ,常规培养方法检出 33株 (4 2 .3% ) ,培养法检测阳性者PCR方法均为阳性。PCR方法较培养法有显著性差异。结论 :PCR方法检测呼吸道流感嗜血杆菌感染比常规方法更快速 ,更简便 ,更敏感。PCR方法检测流感嗜血杆菌有可能成为临床流感嗜血杆菌感染诊断的一种实用、理想的方法。     

Quantification of Helicobacter pylori levels in soil samples from public playgrounds in Spain

Helicobacter pylori are ubiquitous Gram-negative bacteria with a high estimated level of infection in the world populations, but a majority of the infected persons are asymptomatic. This pathogen has been classified by the World Health Organization as a class I carcinogen and recognized as the causal agent of most peptic ulcers and chronic gastritis that might lead to stomach cancer. Although not all the transmission pathways of these bacteria into humans have been properly identified, enough data have suggested that the oral-oral or fecal-oral ones are the main infection routes. Helicobacter pylori have been detected in non-treated water and in drinking water, which suggested that water might be an important infection source. As childhood is the critical period of infection, the aim of the present work was to examine the presence of Helicobacter pylori in soil samples from public playing areas of Spanish parks.     

MGB probe assay for rapid detection of mtDNA11778 mutation in the Chinese LHON patients by real-time PCR

Objective:Leber's hereditary optic neuropathY (LHON)is a maternally inherited degeneration of the optic nerve caused by point mutations of mitochondrial DNA(mtDNA).Many unsolved questions regarding the penetrance and pathophysiological mechanism of LHON demand efficient and reliable mutation testing.This study aims to develop a minor groove binder(MGB) probe assay for rapid detection of mtDNA11778 mutation and heteroplasmy in Chinese LHON patients by real-time polymerase chain reaction(PCR).Methods:Forty-eight patients suspected of having LHON and their maternal relatives underwent a molecular genetic evaluation,with 20 normal individuals as a control group at the same time.A real-time PCR involving two MGB probes was used to detect the mtDNA 11778 mutation and heteroplasmy.A linear standard curve was obtained by pUCmLHONG and pUCmLHONA clones.Results:All 48 LHON patients and their matemal relatives were positive for mtDNA 11778 mutation in our assay,27 heteroplasmic and 21 homoplasmic.Eighteen cases did not show an occurrence of the disease,while 9 developed the disease among the 27 heteroplasmic mutation cases.Eleven did not show an occurrence of the disease,while 10 cases developed the disease among 21 homoplasmic mutation cases.There was a significant difierence in the incidence between the heteroplasmic and the homoplasmic mutation types.The time needed for running a real-time PCR assay was only 80 min.Conclusion:This real-time PCR assay is a rapid,reliable method for mtDNA mutation detection as well as heteroplasmy quantification.Detecting this ratio is very important for predicting phenotypic expression of unaffected carriers.     

利用已知微卫星引物寻找目的DNA

利用聚合酶链式反应(PCR)方法特异性扩增河北省饶阳县大仓鼠(Circetulvs triton)基因组DNA通过琼脂糖凝胶电泳检测以期出现特异性条带,可以进一步回收进行序列分析．     

On-Site Pedagogical Content Knowledge Development

Experiences and reflection have long been regarded as a foundation for pedagogical content knowledge (PCK) development. However, little is known about how experienced teachers develop their PCK via reflection-in-action during their moment-to-moment classroom instruction. Drawing upon data sources including classroom observations, semi-structured interviews and stimulated recall interviews based on lesson videos, this study examined instances when four experienced teachers were found to invent new instructional strategies/representations on the spot during the lesson (referred to as on-site PCK development) in their first attempts at teaching a new topic. The study documented the moment-to-moment experiences of the teachers, including their reconstructed thought processes associated with these instances of on-site PCK development. An explanatory model of a three-step process comprising a stimulus, an integration process and a response was advanced to account for the on-site PCK development observed among the teachers. Three categories of stimulus that triggered on-site PCK development were identified. Factors influencing the integration process and, hence, the resulting response, included teachers’ subject matter knowledge of the new topic, their general pedagogical knowledge and their knowledge of student learning difficulties/prior knowledge related to the new topic. Implications for teacher professional development in terms of how to enhance teachers’ on-site PCK development are discussed.     

Detection of PCV2 DNA by SYBR Green I-based quantitative PCR

We developed an assay for the detection and quantitation of porcine circovirus type 2 (PCV2) with the SYBR Green I-based real-time PCR. The real-time PCR provides a broad dynamic range, detecting from 103 to 1011 copies of DNA per reaction. No cross-reactions were found in specimens containing PCV1. Because of the high sensitivity and specificity of the assay with a relatively rapid and simple procedure, real-time PCR can be used as a routine assay for the clinical diagnosis of PCV2 infection. In this study we applied real-time PCR assay to 80 clinical samples, collected from 40 pigs with postweaning multisystemic wasting syndrome (PMWS) and 40 healthy pigs in comparison with conventional PCR assay. In 56 of 80 samples, PCV2 DNA was de-tected by conventional PCR assay. All samples positive for PCV2 DNA in conventional PCR assay were also positive in real-time assay, and 12 of 24 samples that tested negative for PCV2 DNA in the conventional assay were tested positive in real-time PCR assay. Real-time PCR assay increased the number of samples in which PCV2 was detected by 15%. It is, therefore, considered to be a useful tool for the detection of PCV2.     

Quantitative analysis of a panel of gene expression in prostate cancer——with emphasis on NPY expression analysis

Objective： To investigate molecular alterations associating with prostate carcinoma progression and potentially provide information toward more accurate prognosis/diagnosis. Methods： A set of laser captured microdissected （LCM） specimens from 300 prostate cancer （PCa） patients undergoing radical prostatectomy （RP） were defined. Ten patients representing ＂aggressive＂ PCa, and 10 representing ＂non-aggressive＂ PCa were selected based on prostate-specific antigen （PSA） recurrence, Gleason score, pathological stage and tumor cell differentiation, with matched patient age and race between the two groups. Normal and neoplastic prostate epithelial cells were collected with LCM from frozen tissue slides obtained from the RP specimens. The expressions of a panel of genes, including NPY, PTEN, AR, AMACR, DD3, and GSTP1, were measured by quantitative real-time RT-PCR （TaqMan）, and correlation was analyzed with clinicopathological features. Results： The expressions of AMACR and DD3 were consistently up-regulated in cancer cells compared to benign prostate epithelial cells in all PCa patients, whereas GSTP1 expression was down regulated in each patient. NPY, PTEN and AR exhibited a striking difference in their expression patterns between aggressive and non-aggressive PCas （P=0.0203, 0.0284, and 0.0378, respectively, Wilcoxon rank sum test）. The lower expression of NPY showed association with ＂aggressive＂ PCas based on a larger PCa patient cohort analysis （P=0.0037, univariate generalized linear model （GLM） analysis）. Conclusion： Despite widely noted heterogeneous nature of PCa, gene expression alterations ofAM,4CR, DD3, and GSTP1 in LCM-derived PCa epithelial cells suggest for common underlying mechanisms in the initiation of PCa. Lower NPY expression level is significantly associated with more aggressive clinical behavior of PCa; PTEN and AR may have potential in defining PCa with aggressive clinical behavior. Studies along these lines have potential to define PCa-associated gene expression alterations and likely co-regulation of genes/pathways critical in the biology of PCa onset/progression.     

Quantitative analysis of a panel of gene expression in prostate cancer——with emphasis on NPY expression analysis

Objective: To investigate molecular alterations associating with prostate carcinoma progression and potentially provide information toward more accurate prognosis/diagnosis. Methods: A set of laser captured microdissected (LCM) specimens from 300 prostate cancer (PCa) patients undergoing radical prostatectomy (RP) were defined. Ten patients representing \"aggressive\" PCa, and 10 representing \"non-aggressive\" PCa were selected based on prostate-specific antigen (PSA) recurrence,Gleason score, pathological stage and tumor cell differentiation, with matched patient age and race between the two groups.Normal and neoplastic prostate epithelial cells were collected with LCM from frozen tissue slides obtained from the RP specimens.The expressions of a panel of genes, including NPY, PTEN, AR,AMACR, DD3, and GSTP1, were measured by quantitative real-time RT-PCR (TaqMan), and correlation was analyzed with clinicopathological features. Results: The expressions of AMACR and DD3 were consistently up-regulated in cancer cells compared to benign prostate epithelial cells in all PCa patients, whereas GSTP1 expression was down regulated in each patient. NPY, PTEN and AR exhibited a striking difference in their expression patterns between aggressive and non-aggressive PCas (P=0.0203, 0.0284, and 0.0378, respectively, Wilcoxon rank sum test). The lower expression of NPY showed association with \"aggressive\" PCas based on a larger PCa patient cohort analysis (P=0.0037,univariate generalized linear model (GLM) analysis). Conclusion: Despite widely noted heterogeneous nature of PCa, gene expression alterations of AMACR, DD3, and GSTP1 in LCM-derived PCa epithelial cells suggest for common underlying mechanisms in the initiation of PCa. Lower NPY expression level is significantly associated with more aggressive clinical behavior of PCa; PTEN and AR may have potential in defining PCa with aggressive clinical behavior. Studies along these lines have potential to define PCa-associated gene expression alterations and likely co-regulation of genes/pathways critical in the biology of PCa onset/progression.     

应用实时荧光定量PCR检测子宫内膜癌与癌旁组织中microRNA-93的差异表达状况

为分析microRNA-93在人子宫内膜癌组织和癌旁正常组织中的表达差异状况,初步探讨其在子宫内膜癌中的作用,应用实时荧光定量PCR方法检测32例子宫内膜癌及对照癌旁组织中microRNA-93的表达状况。结果表明,子宫内膜癌组织中microRNA-93的表达量高于癌旁组织（P<0.01）,可见microRNA-93可能作为癌基因参与子宫内膜癌的发生发展过程。     

张家口地区汉族人群3个STR基因座的多态性调查

目的:调查D16S539、D7S820及D13S317三个STR基因座在张家口地区汉族人群的等位基因分布,获得相关群体遗传学参数.方法:用SilverⅢ Multipiex试剂盒复合扩增,聚丙烯凝胶电泳分型,银染显带.结果:108例汉族群体在3个基因座上均各检出7个等位基因,多态性分别为0.7314、0.7371、0.7635;个人识别率依次为0.8912、0.8948、0.9223,累计0.9986.结论:3个STR基因座在本地有较高的遗传多态性,在法医学及人类遗传学方面有较好的应用价值.