铜绿假单胞菌PA0861突变菌株表达谱检测分析

目的 分析铜绿假单胞菌野生型菌株PAO1及PA0861突变菌株基因表达谱,探讨铜绿假单胞菌中GGDEF-EAL基因PA0861的生物学功能. 方法 以野生型铜绿假单胞菌PAO1及PA0861基因转座子插入失活的突变体菌株为材料,利用结晶紫染色方法进行细菌生物膜的表型分析;利用Microarray技术分析其基因表达谱的变化;采用半定量RT-PCR验证Microarray结果. 结果 PA0861突变株生物膜的合成量在4、8、12h均高于PAO1 (P＜0.05),在12h时增加了近1倍;利用Microarray技术筛选出差异表达基因517个,其中319个上调,198个下调.生物分子功能注释和Pathway分类显示这些差异表达基因除涉及代谢过程中基本的物质变化和能量转换外,突变体PA0861中涉及铁载体生物合成基因的表达增高.半定量RT-PCR验结果一致. 结论 铜绿假单胞菌突株PA0861基因表达谱发生改变,其中铁载体生物合成基因高表达,提示c-di-GMP可能通过对铁载体的调控影响细菌生物膜的形成.

疾病体外检测和诊断技术的新发展

从全球范围来看,疾病体外检测、诊断试剂的需求在不断增长,年增长率达到5.5％;我国诊断试剂的市场增长率达到近20％[1].特异、敏感和快速的疾病体外检测和诊断方法是预防和治疗疾病的重要手段和前提.一些新的检测技术和平台正逐渐应用于疾病的检测和诊断,引起了越来越多的关注.随着纳米技术( nanotechnology)和材料的兴起与蓬勃发展,依赖纳米材料和技术的新的疾病诊断方法的研究正在成为疾病检测和诊断的热点;基于纳米材料和技术的新的疾病诊断方法可能成为新一代疾病体外检测和诊断方法而受到世界各国的广泛重视.其他像芯片技术(microarray chip)、微流控技术( microfluidic device)和生物传感器( biosensor)等也开始应用于疾病的体外检测,这些新技术相互结合为新的疾病检测方法研发提供了更多的选择.

AIM To describe a liver cancer = specific gene expression profile and to identify genes that showed alteredexpression between liver cancer tissues and their adjacent nearly normal tissues.METHODS The cDNA probes which were labeled with a-32P dATP were synthesized from total RNA ofliver cancer and adjacent normal tissues and hybridized separately to two identical Atlas human cancer eDNAexpression array membranes containing 588 known genes.RESULTS Autoradiographic results were analyzed by specific Atlas ImageTM (version 1. 0) software.Among the 588 genes analyzed, 18 genes were found up-regulated in cancer, including TFDP2, Aktl, E2F-3etc, and 25 genes were down-regulated in cancer, including TDGF1, BAK, LAR, etc. Expression levels ofgenes that associated with the regulation of cell proliferation, apoptosis, differentiation, cell-cellinteraction, invasion regulators and eytokines altered mostly.CONCLUSION The result obtained from Atlas microarray provides a comprehensive liver cancer-specificexpression profile. The results can lead to the identification of liver cancer-specific biomarkers and may behelpful in early diagnosis and dentifiction of target genes for designing rational therapeutic strategies.

全基因组芯片与目的基因芯片在产前诊断中的应用价值及比较

根据今年9月30日发表在Prenat Diagn杂志上的文章报道认为,运用全基因组微阵列芯片(whole-genome microarray)比目的序列微阵列芯片(targeted microarray)更易检测出孕妇样本中具有临床意义的染色体异常.

Wallerian degeneration is a subject of major interest in neuroscience. A large number of genes are differentially regulated during the distinct stages of Wallerian degeneration: transcription factor activation, immune response, myelin cell differentiation and dedifferentiation. Although gene expression responses in the distal segment of the sciatic nerve after peripheral nerve injury are known, differences in gene expression between the proximal and distal segments remain unclear. In the present study in rats, we used microarrays to analyze changes in gene expression, biological processes and signaling pathways in the proximal and distal segments of sciatic nerves under-going Wallerian degeneration. More than 6,000 genes were differentially expressed and 20 types of expression tendencies were identiifed, mainly between proximal and distal segments at 7-14 days after injury. The differentially expressed genes were those involved in cell differentiation, cytokinesis, neuron differentiation, nerve development and axon regeneration. Furthermore, 11 biological processes were represented, related to responses to stimuli, cell apoptosis, inlfammato-ry response, immune response, signal transduction, protein kinase activity, and cell proliferation. Using real-time quantitative PCR, western blot analysis and immunohistochemistry, microarray data were veriifed for four genes: aquaporin-4, interleukin 1 receptor-like 1, matrix metallopro-teinase-12 and periaxin. Our study identiifes differential gene expression in the proximal and distal segments of a nerve during Wallerian degeneration, analyzes dynamic biological changes of these genes, and provides a useful platform for the detailed study of nerve injury and repair during Wallerian degeneration.

This study investigated the possible involvement of microRNAs in the regulation of genes that participate in peripheral neural regeneration. A microRNA microarray analysis was conducted and 23 microRNAs were identiifed whose expression was signiifcantly changed in rat dorsal root ganglia after sciatic nerve transection. The expression of one of the downregulated microRNAs, microRNA-214, was validated using quantitative reverse transcriptase-PCR. MicroRNA-214 was predicted to target the 3′-untranslated region of Slit-Robo GTPase-activating protein 3. In situ hybridization veriifed that microRNA-214 was located in the cytoplasm of dorsal root ganglia primary neurons and was downregulated following sciatic nerve transection. Moreover, a com-bination of in situ hybridization and immunohistochemistry revealed that microRNA-214 and Slit-Robo GTPase-activating protein 3 were co-localized in dorsal root ganglion primary neu-rons. Western blot analysis suggested that Slit-Robo GTPase-activating protein 3 was upregulated in dorsal root ganglion neurons after sciatic nerve transection. These data demonstrate that mi-croRNA-214 is located and differentially expressed in dorsal root ganglion primary neurons and may participate in regulating the gene expression of Slit-Robo GTPase-activating protein 3 after sciatic nerve transection.

基因芯片技术及其在耳聋研究中的应用

基因芯片又称DNA芯片(DNA chip)、DNA微阵列(DNA microarray)、寡核苷酸阵列(Oligonucleotide array)等.基因芯片技术是融微电子学、生物学、物理学、化学、计算机科学等多学科为一体的新技术.它可以将生物中许多不连续的分析过程,移植到固相的介质芯片,并使其连续化和微型化.

Advancement in microarray technology can revolutionize many aspects of medicine. Microarrays have applications in gene expression profiling, genotyping, mutation analysis, gene identification, and pharmacology. This paper provides a brief review on the use of microarrays in studies of cancer, infectious diseases, chromosome disorders, neurological/mental disorders, and drugs, along with a prospect on its great potential in diagnosis, prognosis and the treatment of human diseases.

皮肤石蜡包埋标本组织芯片制备条件优化及应用评价

组织芯片(tissue chip )又称组织微阵列(tissue microarray,TMA),是将若干石蜡包埋块上的各种典型组织或细胞转移到一个新石蜡块上重新构建微型化高通量组织阵列的方法.1

应用cDNA文库法制备HCV-1诊断芯片探针

目的探讨利用cDNA文库法制备丙型肝炎(HCV-1)诊断基因芯片探针的可行性.方法用限制性内切酶Sau3AⅠ消化HCV 1 a及1b全长cDNA,所得的酶切片段72℃补平加A,AT克隆,PCR初步鉴定,并测序.结果 HCV两个亚型1a、1b的全长cDNA得到57个大小相对一致(200-1000bp)的片段,平均每个亚型约28个,PCR及序列分析表明,所扩增的片段均属于HCV-1的特异基因,可做为HCV-1诊断基因芯片探针.结论利用cDNA文库法收集片段是一种快速、简便制备芯片探针的实用方法.

Oestrogen is essential for maintaining bone mass, and it has been demonstrated to induce osteoblast proliferation and bone formation. In this study, complementary DNA (cDNA) microarrays were used to identify and study the expression of novel genes that may be involved in MC3T3-E1 cells’ response to 17-b estradiol. MC3T3-E1 cells were inoculated in minimum essential media alpha (a-MEM) cell culture supplemented with 17-b estradiol at different concentrations and for different time periods. MC3T3-E1 cells treated with 1028 mol?L21 17-b estradiol for 5 days exhibited the highest proliferation and alkaline phosphatase (ALP) activity;thus, this group was chosen for microarray analysis. The harvested RNA was used for microarray hybridisation and subsequent real-time reverse transcription polymerase chain reaction (RT-PCR) to validate the expression levels for selected genes. The microarray results were analysed using both functional and pathway analysis. In this study, microarray analysis detected 5 403 differentially expressed genes, of which 1 996 genes were upregulated and 3 407 genes were downregulated, 1 553 different functional classifications were identified by gene ontology (GO) analysis and 53 different pathways were involved based on pathway analysis. Among the differentially expressed genes, a portion not previously reported to be associated with the osteoblast response to oestrogen was identified. These findings clearly demonstrate that the expression of genes related to osteoblast proliferation, cell differentiation, collagens and transforming growth factor beta (TGF-b)-related cytokines increases, while the expression of genes related to apoptosis and osteoclast differentiation decreases, following the exposure of MC3T3-E1 cells to a-MEM supplemented with 17-b estradiol. Microarray analysis with functional gene classification is critical for a complete understanding of complementary intracellular processes. This microarray analysis provides large-scale gene expression data that require further confirmatory studies.

Objective: The aim of this study was to reveal the exact changes during the occurrence of breast cancer to explore significant new and promising genes or factors related to this disease.
Methods: We compared the gene expression profiles of breast cancer tissues with its uninvolved normal breast tissues as controls using the cDNA microarray analysis in seven breast cancer patients. Further, one representative gene, named IFI30, was quanti-tatively analyzed by real-time PCR to confirm the result of the cDNA microarray analysis.
Results: A total of 427 genes were identified with significantly differential expression, 221 genes were up-regulated and 206 genes were down-regulated. And the result of cDNA microarray analysis was validated by detection of IFI30 mRNA level changes by real-time PCR. Genes for cell proliferation, cell cycle, cell division, mitosis, apoptosis, and immune response were enriched in the up-regulated genes, while genes for cell adhesion, proteolysis, and transport were significantly enriched in the down-regulated genes in breast cancer tissues compared with normal breast tissues by a gene ontology analysis.
Conclusion: Our present study revealed a range of differentially expressed genes between breast cancer tissues and normal breast tissues, and provide candidate genes for further study focusing on the pathogenesis and new biomarkers for breast cancer.
Copyright ? 2015, Chinese Medical Association Production. Production and hosting by Elsevier B.V. on behalf of KeAi Communications Co., Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).