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超高速细胞分选平台结合cdna microarray技术筛查宫颈癌细胞潜在分子标志物

重庆医科大学学报 2023年第1期48卷 54-60页

作者：陈红香周自华周艳宏新疆维吾尔自治区人民医院妇科乌鲁木齐830001 湖南娄底市中心医院肿瘤科长沙410078 中南大学基础医学院肿瘤研究所长沙410011

目的:通过超高速细胞分选平台结合cdna microarray技术,筛查宫颈癌细胞可能潜在的分子标志物。方法:采用MoFlo XDP型超高速细胞分选平台纯化细胞膜表面表达CD38和不表达CD38的宫颈癌细胞,利用RNAlater技术得到cdna microarray实验所需R... 详细信息

目的:通过超高速细胞分选平台结合cdna microarray技术,筛查宫颈癌细胞可能潜在的分子标志物。方法:采用MoFlo XDP型超高速细胞分选平台纯化细胞膜表面表达CD38和不表达CD38的宫颈癌细胞,利用RNAlater技术得到cdna microarray实验所需RNA,然后进行基因芯片分析。结果:利用MoFlo XDP型超高速细胞分选平台可以获得纯度为99.0%以上的CD38阳性表达宫颈癌细胞。结论:cdna microarray分析发现了RORA、PLIN4、AUTS2、IFITM1等宫颈癌细胞潜在分子标志物,为宫颈癌研究提供了新的技术方法。

关键词：流式细胞术 cdna microarray 宫颈癌分子标志物

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cdna microarray analysis of human keratinocytes cells of patients submitted to chemoradiotherapy and oral photobiomodulation therapy: pilot study

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LASERS IN MEDICAL SCIENCE 2018年第1期33卷 11-18页

作者： Antunes, Heliton S. Wajnberg, Gabriel Pinho, Marcos B. Nogueira Jorge, Natasha Andressa Melo de Moraes, Joyce Luana Stefanoff, Claudio Gustavo Herchenhorn, Daniel Araujo, Carlos M. M. Pais Viegas, Celia Maria Rampini, Mariana P. Dias, Fernando L. de Araujo-Souza, Patricia Savio Passetti, Fabio Ferreira, Carlos G. Inst Nacl Canc INCA Div Clin Res Rua Andre Cavalcante 372 Andar BR-20231050 Rio De Janeiro RJ Brazil Inst Nacl Canc INCA Bioinformat Unit Clin Res Coordinat Rio De Janeiro RJ Brazil Grp Oncol DOr Head & Neck Oncol Grp Rio De Janeiro Brazil INCA Div Radiat Oncol Rio De Janeiro Brazil INCA Head & Neck Surg Div Rio De Janeiro Brazil INCA Program Cellular Biol Rio De Janeiro Brazil Univ Fed Fluminense Dept Immunobiol Niteroi RJ Brazil Fundacao Oswaldo Cruz Fiocruz Inst Oswaldo Cruz Lab Funct Genom & Bioinformat Rio De Janeiro RJ Brazil DOr Inst Res & Educ Rio De Janeiro Brazil

Oral mucositis is an acute toxicity that occurs in patients submitted to chemoradiotherapy to treat head and neck squamous cell carcinoma. In this study, we evaluated differences in gene expression in the keratinocytes of the oral mucosa of patients treated with photobiomodulation therapy and tried to associate the molecular mechanisms with clinical findings. From June 2009 to December 2010, 27 patients were included in a randomized double-blind pilot study. Buccal smears from 13 patients were obtained at days 1 and 10 of chemoradiotherapy, and overall gene expression of samples from both dates were analyzed by complementary DNA (cdna) microarray. In addition, samples from other 14 patients were also collected at D1 and D10 of chemoradiotherapy for subsequent validation of cdna microarray findings by qPCR. The expression array analysis identified 105 upregulated and 60 downregulated genes in our post-treatment samples when compared with controls. Among the upregulated genes with the highest fold change, it was interesting to observe the presence of genes related to keratinocyte differentiation. Among downregulated genes were observed genes related to cytotoxicity and immune response. The results indicate that genes known to be induced during differentiation of human epidermal keratinocytes were upregulated while genes associated with cytotoxicity and immune response were downregulated in the laser group. These results support previous clinical findings indicating that the lower incidence of oral mucositis associated with photobiomodulation therapy might be correlated to the activation of genes involved in keratinocyte differentiation.

关键词： Photobiomodulation therapy (PBM therapy) Oral mucositis Head and neck cancer Radiotherapy Chemotherapy cdna microarray qPCR

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Contrast enhancement for cdna microarray image based on fourth-order moment

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SIGNAL IMAGE AND VIDEO PROCESSING 2018年第6期12卷 1069-1077页

作者： Li, Tiejun Shao, Guifang Sun, Yue Shi, Weiren Chongqing Univ Inst Automat Chongqing 400044 Peoples R China Jimei Univ Sch Informat Engn Xiamen 361021 Peoples R China Xiamen Univ Dept Automat Xiamen 361005 Peoples R China

microarray technology, which can monitor the expression levels of thousands of genes simultaneously, has been widely used in biological experiment. Image processing, as one key step in microarray technology, plays an essential role in microarray analysis. Meanwhile, biological applications require a higher accuracy in each image processing step. However, the low contrast levels of image make it difficult to obtain better processing precision. This paper proposes a fully automatic contrast enhancement (CE) method based on fourth-order moment. Also, a background estimation method is presented to obtain a better microarray image processing result. Comparative results on contrast enhance and gridding revealed that the proposed CE algorithm performs better compared to the adaptive histogram equalization method. Numerous experiments on the Swiss Institute of Bioinformatics (SIB), Joe DeRisi individual (DeRisi), Gene Expression Omnibus (GEO), and Stanford microarray Database (SMD) data sets also indicate that the proposed CE exerts a tremendous effect on gridding, but has nothing to do with segmentation.

关键词： cdna microarray Contrast enhancement Spot Gridding Segmentation

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cdna microarray Analysis of Insulin Resistance-associated Genes in Fructose-fed Rats

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Agricultural Biotechnology 2018年第4期7卷 93-96,99页

作者： Bo WANG Yuming LI Xuyan ZHAO Shaoxi CAO Yanwei ZHANG Teaching Laboratory Management Center Guizhou Normal CollegeGuiyang 550018China Institute of Cardiovascular Disease Pingjin HospitalMedical College of Chinese People s Armed Police ForcesTianjin 300162China Bioengineering Institute Key Laboratory of Biomechanics&Tissue Engineering Ministry of EducationChongqing UniversityChongqing 400044China Guizhou Bioresource Development and Utilization Key Laboratory Guiyang 550018China

This study was conducted to investigate the gene expression in fructose-fed rat skeletal muscle by cdna chip which could provide support to elucidate the molecular mechanisms underlying insulin resistance. The rats were divided into two groups, one of which was normal control and the other was fed with fructose-rich diet. The mRNA was isolated and purified from the skeletal muscle of two groups. The mRNA from two kinds of tissue was reverse transcribed to cdna with Cy3-dUTP and Cy5-dUTP separately to prepare hybridization probes. The mixed probes were hybridized to cdna microarray. The microarray was scanned, analyzed and repeated for two times. Among the total 4 096 tested genes, 140 genes were differently expressed, 62 up-regulated,78 down-regulated, the expression of Ptprd and Gilz and multiple genes of oxidative metabolism is associated in insulin resistance. The differential expression of gene may be related to the pathogenesis of insulin resistance.

关键词： cdna microarray Gene expression profile Insulin resistance Fructose-fed rats

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Sulforaphane-Induced Apoptosis in Human Leukemia HL-60 Cells Through Extrinsic and Intrinsic Signal Pathways and Altering Associated Genes Expression Assayed by cdna microarray

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ENVIRONMENTAL TOXICOLOGY 2017年第1期32卷 311-328页

作者： Shang, Hung-Sheng Shih, Yung-Luen Lee, Ching-Hsiao Hsueh, Shu-Ching Liu, Jia-You Liao, Nien-Chieh Chen, Yung-Liang Huang, Yi-Ping Lu, Hsu-Feng Chung, Jing-Gung Triserv Gen Hosp Div Clin Pathol Natl Def Med Ctr Dept Pathol Taipei Taiwan Fu Jen Catholic Univ Sch Med Taipei Taiwan Shin Kong Wu Ho Su Mem Hosp Dept Pathol & Lab Med Taipei Taiwan Taipei Med Univ Sch Med Lab Sci & Biotechnol Taipei Taiwan Jen Jr Coll Med Nursing & Management Houlong Township Miaoli County Taiwan Cheng Hsin Gen Hosp Dept Clin Pathol Taipei Taiwan Yuanpei Univ Dept Med Lab Sci & Biotechnol Hsinchu Taiwan China Med Univ Dept Physiol Taichung 404 Taiwan Fu Jen Catholic Univ Dept Restaurant Taipei Taiwan Fu Jen Catholic Univ Dept Hotel & Inst Management Taipei Taiwan China Med Univ Dept Biol Sci & Technol Taichung Taiwan Asia Univ Dept Biotechnol Taichung Taiwan

Sulforaphane (SFN), one of the isothiocyanates, is a biologically active compound extracted from cruciferous vegetables, and has been shown to induce cytotoxic effects on many human cancer cells including human leukemia cells. However, the exact molecular mechanism and altered gene expression associated with apoptosis is unclear. In this study, we investigated SFN-induced cytotoxic effects and whether or not they went through cell-cycle arrest and induction of apoptosis and further examined molecular mechanism and altered gene expression in human leukemia HL-60 cells. Cell viability, cellcycle distribution, sub-G1 (apoptosis), reactive oxygen species (ROS) and Ca2+ production, levels of mitochondrial membrane potential (Delta psi(m)), and caspase-3, 28, and 29 activities were assayed by flow cytometry. Apoptosis-associated proteins levels and gene expressions were examined by Western blotting and cdna microarray assays, respectively. Results indicated that SFN decreased viable cells, induced G2/M phase arrest and apoptosis based on sub-G1 phase development. Furthermore, SFN increased ROS and Ca2+ production and decreased the levels of Delta psi(m) and activated caspase-3, -8, and -9 activities in HL-60 cells. SFN significantly upregulated the expression of BAX, Bid, Fas, Fas-L, caspase-8, Endo G, AIF, and cytochrome c, and inhibited the antiapoptotic proteins such as Bcl-x and XIAP, that is associated with apoptosis. We also used cdna microarray to confirm several gene expressions such as caspase -8, -3, -4, -6, and -7 that are affected by SFN. Those results indicated that SFN induced apoptosis in HL-60 cells via Fas- and mitochondria-dependent pathways. (C) 2016 Wiley Periodicals, Inc.

关键词： sulforaphane apoptosis human leukemia HL-60 cells cdna microarray

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A cdna microarray analysis of the molecular control of poplar wood properties

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Journal of Forestry Research 2017年第1期28卷 71-82页

作者： Qinjun Huang Changjun Ding Weixi Zhang Bingyu Zhang Yanguang Chu Dahai Wang Xiaohua Su State Key Laboratory of Tree Genetics and Breeding Research Institute of ForestryChinese Academy of ForestryBeijing 100091China Key Laboratory of Tree Breeding and Cultivation of State Forestry Administration Beijing 100091China

Molecular biological research into wood development and formation has been the focus in recent years, but the pace of discovery of related genes and their functions in the control of wood properties has been slow. The microarray technique--with its advantages of high throughput capacity, sensitivity, and reliability over other tools developed for investigating genes expression patterns-is capable of rapidly assaying thousands of genes. In this study, a cdna microarray prepared from two cdna libraries of developing poplar xylem tissues was used to assay gene expression patterns in immature xylem tissues at different heights from the main stem of Populus deltoides （15 years old）, which was confirmed to have distinct wood properties （microfibrillar angle, woody density） by X-ray. Two hundred seventy-four transcripts with differ- ential expression profles between the chips were screened out, and the individual clones were subjected to 5＇ sequencing. Using bioinformatic analysis, we identified candidate genes that may influence poplar wood properties, many of which belong to various regulatory and signal transduction gene families, such as zinc finger protein transcription factor, DNA-binding transcription factor, ethylene response factors, and so on. The results suggest that these genes may regulate enzymes involved in wood formation. Further work will be performed to clone these genes and determine how they influence poplar wood properties.

关键词： Populus deltoides cdna microarray Expression profile Wood density Microfibfillar angle

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Optimization of Cell-Based cdna microarray Conditions for Gene Functional Studies in HEK293 Cells

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SLAS DISCOVERY 2017年第8期22卷 1053-1059页

作者： Kim, Hi Chul Heo, Jin Yeong Lee, Tae-Kyu Cho, Ssang-Goo Kwon, Yong-Jun Inst Pasteur Korea Seongnam Gyeonggi Do South Korea Konkuk Univ Dept Anim Biotechnol Seoul 05029 South Korea Konkuk Univ Incurable Dis Anim Model & Stem Cell Inst IDASI Seoul 05029 South Korea Ksilink 16 Rue Ankara F-67000 Strasbourg France

Since the cell-based cdna microarray (CBCM) technique has been a useful tool for gain-of-function studies, many investigators have used CBCMs to identify interesting genes. However, this method requires better-established conditions to ensure high reverse transfection efficiency without cross-contamination. Therefore, we optimized CBCM techniques through various means. We determined that Lipofectamine 2000 was the most appropriate transfection reagent by evaluating eight commercialized reagents, and we determined that the most effective concentrations for printing solution constituents were 0.2 M sucrose (to yield a final concentration of 32 mM) and 0.2% gelatin (to yield a final concentration 0.075%). After examining various combinations, we also determined that the best concentrations of cdna and transfection reagent for optimal reverse transfection efficiency were 1.5 mu g/5 mu L of cdna and 5.5 mu L of Lipofectamine 2000. Finally, via a time course, we determined that 72 h was the most effective reaction duration for reverse transfection, and we confirmed the stability of cdna spot activity of CBCMs for various storage periods. In summary, the CBCM conditions that we have identified can provide more effective outcomes for cdna reverse transfection on microarrays.

关键词： cdna microarray cell image-based array reverse transfection image-based screening

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Identification of proteasome subunit beta type 2 associated with deltamethrin detoxification in Drosophila Kc cells by cdna microarray analysis and bioassay analyses

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GENE 2016年第1期582卷 85-93页

作者： Hu, Junli Jiao, Dongxu Xu, Qin Ying, Xiaoli Liu, Wei Chi, Qingping Ye, Yuting Li, Xueyu Cheng, Luogen Nanjing Normal Univ Coll Life Sci Jiangsu Key Lab Biodivers & Biotechnol Nanjing 210023 Jiangsu Peoples R China Nanjing Normal Univ Coll Energy & Mech Engn Nanjing 210042 Jiangsu Peoples R China

Insecticide deltamethrin resistance has presented a difficult obstacle for pest control and the resistance development is complex and associated with many genes. To better understand the possible molecular mechanisms involved in DM stress, in this study, cdna microarray analysis was employed. 448 differentially expressed genes with at least a 2-fold expression difference were identified in Drosophila cells after DM exposure. Moreover, some genes were confirmed with qPCR, which yielded results consistent with the microarray analysis. Three members of the ubiquitin-proteasome system were significantly elevated in DM-stressed cells, suggesting that the ubiquitin-proteasome pathway may play an important role in DM detoxification. The proteasome beta2 subunit (Prosbeta2) is a member of 20S proteasome subunit family, which forms the proteolytic core of 26S proteasome. Whether Prosbeta2 participates in DM detoxification requires further study. RNAi and heterologous expression were conducted to investigate the contribution of Prosbeta2 in DM detoxification. The results revealed Prosbeta2 knockdown significantly reduce the level of DM detoxification in RNAi-treated cells after 48 h. Overexpression of Prosbeta2 increased cellular viability. These detoxification results represent the first evidence that Prosbeta2 plays a role in the detoxification of DM, which may provide new idea and target for studying the molecular mechanisms of insect resistance. (C) 2016 Elsevier B.V. All rights reserved.

关键词： Deltamethrin resistance cdna microarray RNAi Heterologous expression Prosbeta2 Overexpression Detoxification

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Comparative transcript profiling of gene expression between self-incompatible and self-compatible mandarins by suppression subtractive hybridization and cdna microarray

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MOLECULAR BREEDING 2015年第1期35卷 1-15页

作者： Miao, Hongxia Ye, Zixing Hu, Guibing Qin, Yonghua South China Agr Univ Coll Hort State Key Lab Conservat & Utilizat Subtrop Agrobi Key Lab Biol & Genet Improvement Hort CropsMinis Guangzhou 510642 Guangdong Peoples R China Chinese Acad Trop Agr Sci Inst Trop Biosci & Biotechnol Minist Agr Key Lab Trop Crop Biosci & Biotechnol Haikou 571101 Hainan Peoples R China

Self-incompatibility (SI) is an important trait of Citrus plants that is exploited by farmers to produce seedless fruit. However, the molecular mechanism of SI in Citrus is not well understood. Wuzishatangju (Citrus reticulata Blanco) (SI) is an excellent seedless cultivar selected from a seedy Shatangju cultivar (self-compatible, SC) through spontaneous bud mutation. The two cultivars are therefore excellent materials for studying the mechanisms of SI and/or SC in Citrus. In this study, an integrative strategy combining eight suppression subtractive hybridization libraries with cdna microarray was used to study the molecular mechanisms that differ between Wuzishatangju and Shatangju (control) mandarins. A custom microarray screen resulted in a total of 1,830 up-or down-regulated clones (false discovery rate <0.05 and a fold change >= 2) obtained from 9,810 positive clones. The expression of genes involved in embryonic development, ubiquitination pathway, Ca2+-signaling pathway, gibberellins, and auxin was significantly upregulated in SI Wuzishatangju compared with SC Shatangju mandarin. The microarray analysis suggested that the ubiquitin-mediated proteolysis pathway might be involved in the SI reaction of Wuzishatangju. Additionally, our research highlighted some main genes (mitogen-activated protein kinase, SI S1 family protein, ubiquitin-conjugating factor E4-like, auxin transporter protein 1, and gibberellin receptor) that participate in the SI reaction of Wuzishatangju and could be beneficial for understanding the evolution of SI systems and for breeding seedless citrus fruits in the future.

关键词： Wuzishatangju (Citrus reticulata Blanco) Self-incompatibility Suppression subtractive hybridization (SSH) cdna microarray Ubiquitin pathway

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Identification of Asian Pear Cultivars S-genotypes by Self-Incompatibility Genes (S-RNase) cdna microarray

Identification of Asian Pear Cultivars S-genotypes by Self-I...

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The Second Asian Horticultural Congress

作者： J.Nan T.Xiaofeng T.Hui School of Packing and Material Engineering Hunan University of Technology Key Laboratory of Cultivation and Protection for Non-wood Forest of Ministry of education the Key Laboratory of Non-wood Forest Product of Forestry MinistryCentral-south University of Forestry and Technology Key Laboratory of Cultivation and Protection for Non-wood Forest of Ministry of Education the Key Laboratory of Non-wood Forest Product of Forestry MinistryCentral-south University of Forestry and Technology

Pears(Pyrus)are gametophytic self-incompatible fruit trees grown *** in different S-genotypes must be present locally to ensure normal pollination and fertilization for high and stable yields as well as successful ***... 详细信息

关键词： cdna microarray S-RNase Identification of Asian Pear Cultivars S-genotypes by Self-Incompatibility Genes

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