Integrated computer system for regulating eukaryotic gene expression

The review describes several modules of the GeneExpress integrated computer system concerning the regulation of gene expression in eukaryotes. Approaches to the presentation of experimental data in databases are considered. The employment of GeneExpress in computer analysis and modeling of the organization and function of genetic systems is illustrated with examples. GeneExpress is available at http://wwwmgs.bionet.nsc.ru/mgs/gnw/.     

An Integrated Computer System for Studying the Regulation of Eukaryotic Gene Expression

The review describes several modules of the GeneExpress integrated computer system concerning the regulation of gene expression in eukaryotes. Approaches to the presentation of experimental data in databases are considered. The employment of GeneExpress in computer analysis and modeling of the organization and function of genetic systems is illustrated with examples. GeneExpress is available at http://wwwmgs.bionet.nsc.ru/mgs/gnw/.     

Nucleosome formation potential of eukaryotic DNA: calculation and promoters analysis.

MOTIVATION: A rapid growth in the number of genes with known sequences calls for developing automated tools for their classification and analysis. It became clear that nucleosome packaging of eukaryotic DNA is very important for gene functioning. Automated computer tools for characterization of nucleosome packaging density could be useful for studying of gene regulation and genome annotation. RESULTS: A program for constructing nucleosome formation potential profiles of eukaryotic DNA sequences was developed. Nucleosome packaging density was analyzed for different functional types of human promoters. It was found that in promoters of tissue-specific genes, the nucleosome formation potential was essentially higher than in genes expressed in many tissues, or housekeeping genes. Hence, capability of nucleosome positioning in the promoter region may serve as a factor regulating gene expression. AVAILABILITY: The program for nucleosome sites recognition is included into the GeneExpress system; section 'DNA Nucleosomal Organization', http://wwwmgs.bionet.nsc.ru/mgs/programs/recon/.     

Recognition of eukaryotic promoters using a genetic algorithm based on iterative discriminant analysis

A new approach to recognizing promoter regions of eukaryotic genes is proposed and illustrated by an example of Drosophila melanogaster. The essence of its novelty is in realizing the genetic algorithm to search for optimal partition of a promoter region into local nonoverlapping fragments and selection of the most significant dinucleotide frequencies for the fragments obtained. The method developed was applied to recognizing TATA-containing (TATA+) and DPE-containing (DPE+) promoters of Drosophila melanogaster genes. The program for promoter recognition is included into the GeneExpress system, section RegScan (http://wwwmgs.bionet.nsc.ru/mgs/programs/proga/).     

Dragon Plant Biology Explorer. A text-mining tool for integrating associations between genetic and biochemical entities with genome annotation and biochemical terms lists

We introduce a tool for text mining, Dragon Plant Biology Explorer (DPBE) that integrates information on Arabidopsis (Arabidopsis thaliana) genes with their functions, based on gene ontologies and biochemical entity vocabularies, and presents the associations as interactive networks. The associations are based on (1) user-provided PubMed abstracts; (2) a list of Arabidopsis genes compiled by The Arabidopsis Information Resource; (3) user-defined combinations of four vocabulary lists based on the ones developed by the general, plant, and Arabidopsis GO consortia; and (4) three lists developed here based on metabolic pathways, enzymes, and metabolites derived from AraCyc, BRENDA, and other metabolism databases. We demonstrate how various combinations can be applied to fields of (1) gene function and gene interaction analyses, (2) plant development, (3) biochemistry and metabolism, and (4) pharmacology of bioactive compounds. Furthermore, we show the suitability of DPBE for systems approaches by integration with "omics" platform outputs. Using a list of abiotic stress-related genes identified by microarray experiments, we show how this tool can be used to rapidly build an information base on the previously reported relationships. This tool complements the existing biological resources for systems biology by identifying potentially novel associations using text analysis between cellular entities based on genome annotation terms. Thus, it allows researchers to efficiently summarize existing information for a group of genes or pathways, so as to make better informed choices for designing validation experiments. Last, DPBE can be helpful for beginning researchers and graduate students to summarize vast information in an unfamiliar area. DPBE is freely available for academic and nonprofit users at http://research.i2r.a-star.edu.sg/DRAGON/ME2/.     

A database of recombinant viruses and recombinant viral vectors available from the RIKEN DNA bank

BACKGROUND: Viral vectors are required as gene-delivery systems for gene therapy and basic research. Recombinant adenoviruses (rAds) expressing genes of interest are being developed as research tools and many studies in vitro and in vivo have already been performed with such rAds. METHODS: Shuttle vectors for rAds were constructed with full-length cDNAs and rAds were generated in HEK293 cells by the COS-TPC method. The rAds and shuttle vectors were developed by the Japanese research community and deposited in the RIKEN DNA Bank (RDB; http://www.brc.riken.jp/lab/dna/en/) for distribution to the scientific community. The Recombinant Virus Database (RVD; http://www.brc.riken.jp/lab/dna/rvd/) was established at the RIKEN BioResource Center (BRC) in Japan as the source of information about and distribution of the various resources. RESULTS: The RIKEN BRC is releasing more than 300 recombinant viruses (RVs) and 500 shuttle vectors, as well as all related information, which is included in a newly established database, the RVD. The RVD consists of (i) information about the RVs, the inserted cDNAs and the shuttle vectors; (ii) data about sequence-tagged sites (STSs) that are markers of viral DNAs; and (iii) experimental protocols for the use of RVs. CONCLUSIONS: The new database and available resources should be very useful to scientists who are studying human gene therapy and performing related basic research. It is a web-interfaced flat-file database that can be accessed through the internet. Moreover, all of the resources deposited in the RDB, which is a public facility in Japan, are available to researchers around the world.     

Systems for the removal of a selection marker and their combination with a positive marker

Many systems have been developed for the removal of a selection marker in order to generate marker-free transgenic plants. These systems consist of (1) a site-specific recombination system (Cre/lox) or a phage-attachment region (attP) to remove the selectable marker gene and (2) a transposable element system (Ac) or a co-transformation system to segregate the gene of interest from the selectable marker gene. Overall, the process is more time-consuming than conventional transformation methods because two rounds of transformation - two steps of regeneration or sexual crossings - are required to obtain the desired transgenic plants. Recently, removal systems combined with a positive marker, denoted as MAT vectors, have been developed to save time and effort by generating marker-free transgenic plants through a single-step transformation. We summarize here the transformation procedures using these systems and discuss their feasibility for practical use.     

Resource and hardware options for microarray-based experimentation.

DNA microarray technology permits the study of biological systems and processes on a genome-wide scale. Arrays based on cDNA clones, oligonucleotides and genomic clones have been developed for investigations of gene expression, genetic analysis and genomic changes associated with disease. Over the past 3-4 years, microarrays have become more widely available to the research community. This has occurred through increased commercial availability of custom and generic arrays and the development of robotic equipment that has enabled array printing and analysis facilities to be established in academic research institutions. This brief review examines the public and commercial resources, the microarray fabrication and data capture and analysis equipment currently available to the user.     

Hormone-mediated gene regulation and bioinformatics: learning one from the other

The ability to manage the constantly growing clinically relevant information in genetics available on the internet is becoming crucial in medical practice. Therefore, training students in teaching environments that develop bioinformatics skills is a particular challenge to medical schools. We present here an instructional approach that potentiates learning of hormone/vitamin mechanisms of action in gene regulation with the acquisition and practice of bioinformatics skills. The activity is integrated within the study of the Endocrine System module. Given a nucleotide sequence of a hormone or vitamin-response element, students use internet databases and tools to find the gene to which it belongs. Subsequently, students search how the corresponding hormone/vitamin influences the expression of that particular gene and how a dysfunctional interaction might cause disease. This activity was presented for four consecutive years to cohorts of 50-60 students/year enrolled in the 2(nd) year of the medical degree. 90% of the students developed a better understanding of the usefulness of bioinformatics and 98% intend to use web-based resources in the future. Since hormones and vitamins regulate genes of all body organ systems, this activity successfully integrates the whole body physiology of the medical curriculum.     

基因药物纳米给药系统的设计与研究进展

基因药物的传递面临着体内外稳定性差、缺乏靶向性、难入胞、在细胞内难以释放等一系列障碍和挑战。因此,要实现基因药物在 体内有效传递需构建能克服这些障碍的药物传递系统。随着材料科学和纳米科技的发展,大量新型的纳米载体已被用于基因药物的传递。 综述目前基因药物传递所面临的障碍和挑战,基因药物纳米给药系统的设计思路及研究进展。     

深海白色侧齿霉遗传转化体系建立及渗透压调控相关的EaSHO1基因功能

海洋中具有丰富的动植物及微生物资源,海洋真菌是其重要组成之一。我们前期的研究发现一株深海真菌白色侧齿霉Engyodontium album能产生具有抑菌活性的次级代谢产物engyodontiumin A,该化合物能抑制黑曲霉、金黄色葡萄球菌及创伤弧菌等病原菌的生长,是一种潜在的海洋源抗菌药物。目前,该菌遗传转化体系尚未建立,不利于开展次级代谢产物合成调控机制及其他功能基因研究。本研究成功制备了深海白色侧齿霉菌的原生质体,建立了借助聚乙二醇3350介导的原生质体转化体系,并将pCT74-sGFP载体成功导入白色侧齿霉的原生质体中,结果显示外源GFP能稳定表达。此外,为了明确白色侧齿霉菌是否能够开展基因敲除研究,通过氨基酸序列同源比对,我们选取酵母高渗甘油信号途径中的同源基因EaSHO1进行初步探究。利用同源重组的方法成功将目的基因EaSHO1的开放阅读框(ORF)替换成潮霉素磷酸转移酶基因(HPH),由此获得EaSHO1基因敲除突变体,并对突变体进行Southern杂交验证及初步的表型分析。结果表明,EaSHO1缺失不影响白色侧齿霉菌的营养生长及对高盐胁迫的响应,亚细胞定位结果显示EaS...     

Targeted vectors for gene therapy of cancer and retroviral infections

Gene therapy has developed to a technology which rapidly moved from the laboratory bench to the bedside in the clinic. This implies safe, efficient and targeted gene transfer systems for suitable application to the patient. Beside the development of such gene transfer vectors of viral or nonviral origin, improvement of cell type specific and inducible gene expression is pivotal for successful gene therapy leading to targeted gene action. Numerous gene therapy approaches for treatment of cancer and retroviral infections utilize cell type specific and/or regulatable promoter and enhancer sequences for the selective expression of therapeutic genes in the desired cell populations and tissues. In this article the recent developments and the potential of expression targeting are reviewed for gene therapy approaches of cancer and retroviral infections.     

BCDB - A database for breast cancer research and information

In pursuit of a better updated source including 'omics' information for breast cancer, Breast Cancer Database (BCDB) has been developed to provide the researcher with the quick overview of the Breast cancer disease and other relevant information. This database comprises of myriad of information about genes involved in breast cancer, its functions and drug molecules which are currently being used in the treatment of breast cancer. The data available in BCDB is retrieved from the biomedical research literature. It facilitates the user to search information on gene, its location in chromosome, functions and its importance in cancer diseases. Broadly, this can be queried by giving gene name, protein name and drug name. This database is platform independent, user friendly and freely accessible through internet. The data present in BCDB is directly linked to other on-line resources such as NCBI, PDB and PubMed. Hence, it can act as a complete web resource comprising gene sequences, drug structures and literature information related to breast cancer, which is not available in any other breast cancer database. AVAILABILITY: The database is freely available at http://122.165.25.137/bioinfo/breastcancerdb/     

A Grid-based solution for management and analysis of microarrays in distributed experiments

Several systems have been presented in the last years in order to manage the complexity of large microarray experiments. Although good results have been achieved, most systems tend to lack in one or more fields. A Grid based approach may provide a shared, standardized and reliable solution for storage and analysis of biological data, in order to maximize the results of experimental efforts. A Grid framework has been therefore adopted due to the necessity of remotely accessing large amounts of distributed data as well as to scale computational performances for terabyte datasets. Two different biological studies have been planned in order to highlight the benefits that can emerge from our Grid based platform. The described environment relies on storage services and computational services provided by the gLite Grid middleware. The Grid environment is also able to exploit the added value of metadata in order to let users better classify and search experiments. A state-of-art Grid portal has been implemented in order to hide the complexity of framework from end users and to make them able to easily access available services and data. The functional architecture of the portal is described. As a first test of the system performances, a gene expression analysis has been performed on a dataset of Affymetrix GeneChip Rat Expression Array RAE230A, from the ArrayExpress database. The sequence of analysis includes three steps: (i) group opening and image set uploading, (ii) normalization, and (iii) model based gene expression (based on PM/MM difference model). Two different Linux versions (sequential and parallel) of the dChip software have been developed to implement the analysis and have been tested on a cluster. From results, it emerges that the parallelization of the analysis process and the execution of parallel jobs on distributed computational resources actually improve the performances. Moreover, the Grid environment have been tested both against the possibility of uploading and accessing distributed datasets through the Grid middleware and against its ability in managing the execution of jobs on distributed computational resources. Results from the Grid test will be discussed in a further paper.     

基于CRISPR/Cas系统的多重基因编辑与调控技术

规律成簇的间隔短回文重复序列(clustered regularly interspaced short palindromic repeats, CRISPR)及其相关Cas蛋白所构建的CRISPR/Cas系统是古细菌或细菌中特有的一种获得性免疫系统。研究人员将其开发成基因编辑工具之后,凭借其高效、精准和通用性强等优点迅速成为合成生物学领域的热门研究方向,在生命科学、生物工程技术、食品科学及农作物育种等多个领域引发了革命性的影响。目前基于CRISPR/Cas系统单基因编辑与调控技术日益完善,但在多重基因编辑和调控方面仍存在挑战。本文聚焦基于CRISPR/Cas系统的多重基因编辑与调控技术开发及应用,针对单个细胞内实现多位点基因编辑或调控和细胞群体内实现多位点基因编辑或调控技术,依据作用原理对其进行了系统总结和阐述,包括基于CRISPR/Cas系统的双链断裂、单链断裂以及多重基因调控技术等。这些工作丰富了多重基因编辑与调控的工具,为CRISPR/Cas系统在多领域的应用作出了贡献。     

VCre/VloxP and SCre/SloxP: new site-specific recombination systems for genome engineering

We developed two new site-specific recombination systems named VCre/VloxP and SCre/SloxP for genome engineering. Their recognition sites are different from Cre recognition sites because VCre and SCre recombinases share less protein similarity with Cre, even though the basic 13-8-13 structures of their recognition sites are identical. Mutant VloxP and SloxP, which have the same uses as mutant loxP, were also developed. VCre/VloxP and SCre/SloxP in combination with Cre/loxP and Flp/FRT systems can serve as powerful tools for genome engineering, especially when used to genetically modify both alleles of a single gene in mouse and human cells.