Array-Based Gene Expression Analysis - Illumina, Inc.
Data Sheet: Gene Expression Array-Based Gene Expression Analysis Expression profiling products tailored for a variety of genetic research applications. Comprehensive Coverage: Carefully selected array content for discovery of any mRNA Multiple Species Expression profiling solutions for human, mouse, and rat Overview Illumina leads the genetic analysis industry with innovative, flexible products designed for a broad range of genetic research. Its RNA expression profiling assays are performed on Illumina’s BeadArray technology-based systems (Figure 1), providing comprehensive gene coverage on a wide range of sample types. Specifically, Illumina’s Direct Hybridization Whole-Genome Expression assays can be used for analyzing intact RNA samples, while its DASL (cDNA-mediated Annealing, Selection, Extension, and Ligation) assays enable analysis of difficult or degraded RNA samples, such as formalinfixed, paraffin-embedded (FFPE) tissues. These proven assays offer highly accurate data, streamlined workflows, and multi-sample formats for high-throughput multiplex gene expression profiling. Data generated with either of these assays can be visualized and analyzed with Illumina’s GenomeStudio data analysis software. The performance-optimized tools and user-friendly graphical interface of GenomeStudio software allows researchers to quickly and easily convert data into meaningful results. Application Broad Range of Products: Supports discovery, screening, and profiling FFPE samples Profiling Technology High-Quality Data: Rigorously tested assays with internal controls on each array Figure 1: Gene Expression Profiling and Validation Options Product Highlights FFPE / Blood BeadArray Gene Expression BeadChips Whole-Genome DASL-HT Software/Data Analysis Illumina array-based technologies support a broad range of RNA expression profiling products for a variety of applications. highest sample throughput (up to 216 arrays at a time), researchers can automate BeadChip loading and scanning with the AutoLoader, AutoLoader2, or AutoLoader 2.x. Comprehensive Array Content Each probe location (address) and sequence combination on Illumina Expression BeadChips was carefully selected bioinformatically. Gene-specific probes were designed using a multi-step algorithm to optimize several parameters, including: Lack of similarity to other genes Absence of highly repeated sequence in the genome Sequence complexity Self-complementarity for hairpin structure prediction Illumina Expression Arrays Illumina Whole-Genome Gene Expression BeadChips consist of oligonucleotides immobilized to beads held in microwells on the surface of an array substrate (Figure 2). Data quality and reproducibility are supported in part by the high level of bead-type redundancy (up to an average of 30 beads per probe) on every array. After randomly distributing the beads across the substrate surface, 29-mer address sequences present on each bead are used for a hybridization-based procedure to map the array, identifying the location of each bead. This final process also validates the hybridization performance of every bead on every BeadChip, ensuring 100% array quality control. Illumina Expression BeadChip arrays are arranged in a multi-sample format for higher throughput and reduced sample-to-sample variability. Labeled sample cRNA are detected by hybridization to 50-mer probes on the BeadChip. After washing and staining steps, BeadChips are scanned on a HiScan SQ, iScan, or BeadArray Reader. For the Melting temperature for hybridization uniformity Distance from 3’ end of the transcript Illumina expression array solutions include complete kits for human, mouse, and rat whole-genome expression applications, and feature up-to-date content selected from widely used databases such as the National Center for Biotechnology Information Reference Sequence (NCBI RefSeq1) and more specialized sources (Table 1). Exceptional Performance Illumina has compiled performance data for all Expression BeadChips. Reproducibility has been demonstrated by high concordance between technical replicates. Exceptional performance specifications for sensitivity, dynamic range, and fold-change detection precision maximize the impact of differential expression analysis (Table 2).
Data Sheet: Gene Expression Table 1: Expression BeadChip Content Probes Description Human HT-12 v4.0* Mouse WG-6 v2.0 Mouse Ref8 v2.0 Rat Ref-12 Human WG DASL HT* 12-sample 6-sample 8-sample 12-sample 12-sample 27,253 RefSeq Content NM Coding transcript, well-established annotation 28,688 26,766 24,854 6,277 XM Coding transcript, provisional annotation 11,121 6,856 796 15,983 426 NR Non-coding transcript, well-established annotation 1,752 56 47 1 1,580 XR Non-coding transcript, provisional annotation Source RefSeq source release 2,209 Human RefSeq Rel 38 Mouse RefSeq Rel 22 12 26 Rat RefSeq Rel 16 Human RefSeq Rel 38 Supplementary Content UniGene Experimentally confirmed mRNA sequences that align to EST clusters RIKEN FANTOM2 Exemplar protein-coding sequences from the RIKEN FANTOM2 database 5,659 RefSeq Release 5 Transcripts with NM and XM annotation in RefSeq Release 5 (Build 33.1) 3,573 MEEBO Probes to transcripts that do not align with 100% accuracy to RefSeq, but are confirmed as valid mRNA mapping to clusters in Expressed Sequence Tag databases6 2,371 Total 3,461 250 47,231 45,281 25,697 22,523 29,285 * 99.98% of the bead types are present on any HumanHT-12 array Direct Hybridization Assay HumanHT-12 Expression BeadChip The Direct Hybridization Whole-Gene Expression assay offers the highest multiplexing capabilities for whole-genome gene expression, simultaneously profiling more than 47,000 transcripts. By combining the most up-to-date genomic content with low-cost, high-throughput array processing, Illumina Expression BeadChips efficiently and economically deliver high-quality data for gene expression studies of any size. They are ideal for applications such as differential expression analysis, disease classification, pathway analysis, and expressionbased quantitative trait loci (eQTL) studies. The new HumanHT-12 v4.0 Expression BeadChip supports highly efficient human whole-genome gene expression profiling studies with updated content and industry-leading pricing. Its high-value content provides genome-wide transcriptional coverage of wellcharacterized genes, gene candidates, and splice variants, with a significant portion targeting well-established sequences supported by peer-reviewed literature. Figure 2: Direct Hyb Gene Expression Profiling Bead Design Biotin Labeled cRNA Each array on this BeadChip targets more than 47,000 probes. Illumina guarantees that 99.98% of the bead types will be present on any given HumanHT-12 array. This means up to five probes may be represented with only 0, 1, or 2 copies on each HumanHT-12 array. Probes were designed to cover content from NCBI RefSeq Release 38 (November 7, 2009), as well as legacy UniGene content. The 12-sample format facilitates large-scale gene expression applications. With the HumanHT-12 BeadChip, expression information can be easily incorporated into Infinium assay-based genome-wide association studies (GWAS) or methylation studies. MouseWG-6 and MouseRef-8 Expression BeadChips Address Probe Illumina array-based technologies support a broad array of RNA expression profiling products for a variety of applications. The MouseWG-6 v2.0 and MouseRef-8 v2.0 Expression BeadChips are Illumina’s genome-wide gene expression profiling solutions for researchers using the mouse as their model organism. The MouseRef-8 BeadChip features recent content derived from the NCBI RefSeq database. In addition to RefSeq content, the MouseWG-6 BeadChip is supplemented with probes that target the Mouse Exonic Evidence Based Oligonucleotide (MEEBO) set2, as well as exemplar proteincoding sequences described in the RIKEN FANTOM24-6 database3–5.
Data Sheet: Gene Expression Table 2: Direct Hybridization Gene Expression Assay Product Specifications Parameter Specification Probe-Length 50-mer gene-specific probe, plus 29-mer address sequence Sensitivity 1:250,000 Dynamic Range 3 logs Detectable Fold Change 1.35 fold Reproducibility CV 10% Input RNA Required 50–500 ng Expression Array Data Analysis Illumina Expression BeadChips are part of a complete gene expression solution that includes instrumentation, software, and reagent kits. Data analysis is straightforward, since known biologically relevant transcripts are annotated from well-curated databases and probes are designed and validated by Illumina scientists. Illumina’s GenomeStudio Gene Expression Module (Figure 4) enables simplified data management for hierarchical organization of samples, groups, group sets, and all associated project analysis. It offers probelevel and gene-level statistical analysis tools for differential analysis, heat map visualization, and clustering. Figure 3: RNA Profiling with the Whole-Genome DASL HT Assay Researchers can profile from six (MouseWG-6) to eight samples (MouseRef-8) in parallel on a single BeadChip, dramatically increasing throughput while decreasing experimental variability. Illumina’s 100% hybridization-based QC on every probe ensures that Mouse Expression BeadChips deliver the industry’s best performance and reproducibility. Total RNA b b P1 RatRef-12 Expression BeadChip The RatRef-12 Expression BeadChip allows researchers to generate genome-wide expression profiles for twelve samples in parallel on a single BeadChip, with outstanding performance and excellent throughput. BeadChip content (22,523 probes per array) was selected primarily from the NCBI RefSeq database. Low sample input and high-quality data with 100% QC on every feature make the RatRef-12 Expression BeadChip the most comprehensive solution currently available to researchers who are using the rat as a model organism. Whole-Genome DASL HT Assay The Human Whole-Genome DASL HT assay is the only integrated system optimized to produce genome-wide expression profiles from low-abundance or partially degraded human RNA samples, especially those from formalin-fixed, paraffin-embedded (FFPE) tissues. It streamlines the discovery process, allowing researchers to identify new biomarkers by generating expression profiles from archived samples and correlating those profiles with known clinical outcomes. Competing methods for analyzing FFPE samples are based on lightly tested protocols from multiple vendors, which are often both costly and unreliable. Illumina’s Whole-Genome DASL HT assay overcomes these limitations through an innovative combination of two proven assays for genome-wide, highly-sensitive expression profiling. The assay combines the unique PCR and labeling steps from Illumina’s proven DASL Assay with the whole-genome probe set of Illumina’s Direct Hybridization assay (Figure 3). This greatly increases the DASL assay target set, which consists of well-characterized NCBI RefSeq (RefSeq Build 36.2, Release 38) genes, while retaining the ability to accurately profile partially-degraded RNA samples. The Whole-Genome DASL HT assay provides a high-multiplex, low-cost array solution for the simultaneous profiling of over 29,000 transcripts. Highly reproducible expression profiles (r2 0.97) can be produced from as little as 10–100 ng total RNA from fresh-frozen tissue or 50–200 ng total RNA from FFPE samples. P1 P2’ cDNA synthesis Query oligo (DAP) annealing, extension, and ligation P2 PCR with common primers /WV Product captured by hybridization to BeadChip Illumina array-based technologies support a broad array of RNA expression profiling products for a variety of applications. Summary Illumina’s BeadArray-based technology enables cost-effective expression profiling solutions for human, mouse, and rat. Multi-sample BeadChips increase throughput, supporting streamlined experiments. These BeadChips contain comprehensive, up-to-date content derived from several important sources. The DASL assay supports expression profiling from FFPE or RNA-limited samples, and is available as a whole-genome gene expression BeadChip on the BeadArray platform. For any experimental design, Illumina products provide the fastest and most cost-effective path to discoveries and publication.
Data Sheet: Gene Expression Figure 4: GenomeStudio Gene Expression Module The GenomeStudio software interface (left) provides a flexible graphical interface for data and controls display. GenomeStudio software contains powerful built-in data display tools, such as line graphs, tables, and heat maps (right) for expression analysis. References 1. For more information about NCBI RefSeq, please go to ftp://ftp.ncbi.nih.gov/refseq/release/ 2. For more information about MEEBO, please go to http://www.arrays.ucsf.edu/archive/meebo.html 3. For more information about RIKEN FANTOM 2, please go to http://fantom2.gsc.riken.jp/ 4. To obtain FANTOM 2 clones, please go to http://www.dnaform.jp/index e.html 5. The FANTOM Consortium and The RIKEN Genome Exploration Research Group Phase I and II Team (2002) Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs. Nature 420: 563–573. 6. Kuhn K, et al. (2004) A novel , high-performance random array platform for quantitative gene expression profiling. Genome Res 14:2347–2356. 7. Goring HH, et al. (2007) Discovery of Expression Qtls Using Large-Scale Transcriptional Profiling in Human Lymphocytes. Nat Genet 39: 1208–1216. 8. Stranger BE, et al. (2007) Relative Impact of Nucleotide and Copy Number Variation on Gene Expression Phenotypes. Science 315: 848–853. 9. Weinsheimer S, et al. (2007) Integration of Expression Profiles and Genetic Mapping Data to Identify Candidate Genes in Intracranial Aneurysm. Physio Genomics 32: 45–57. 10. Idaghdour Y, et al. (2008) A Genome-Wide Gene Expression Signature of Environmental Geography in Leukocytes of Moroccan Amazighs. PLoS Genet. 4: e1000052. 11. Mutarelli M, et al. (2008) Time-Course Analysis of Genome-Wide Gene Expression Data from Hormone-Responsive Human Breast Cancer Cells. BMC Bioinformatics 9 Suppl 2: S12. 12. Integrating Gene Expression Analysis into Genome-Wide Association Studies (PDF) note integrating expression analysis.pdf 13. RNA Profiling with the DASL Assay Tech Bulletin (PDF) techbulletin dasl.pdf 14. April C, et al. (2009) Whole-Genome Gene Expression Profiling of FormalinFixed, Paraffin-Embedded Tissue Samples. PLoS ONE 4 (12): e8162.
Data Sheet: Gene Expression Ordering Information Product Samples Catalog No. Human Expression BeadChips HumanHT-12 v4.0 Expression BeadChip Kit 72 12 samples per BeadChip 47,000 human targets per sample Includes hybridization buffers, wash buffers, and wash trays Human Whole-Genome DASL HT Assay Kit 12 samples per BeadChip 29,000 human targets per sample Includes reagents for amplifying, hybridizing, washing, and processing BD-103-0604 24 BD-103-0204 24 DA-905-0024 or DA-905-1024 96 DA-905-0096 or DA-905-1096 Mouse Expression BeadChips MouseWG-6 v2.0 Expression BeadChip Kit 12 6 samples per BeadChip 45,000 mouse targets per sample Includes hybridization buffers, wash buffers, and wash trays MouseRef-8 v2.0 Expression BeadChip Kit 36 16 8 samples per BeadChip 25,000 mouse targets per sample Includes hybridization buffers, wash buffers, and wash trays 48 BD-201-0202 BD-201-0602 BD-202-0202 BD-202-0602 Rat Expression BeadChips RatRef-12 Expression BeadChip Kit 24 12 samples per BeadChip 22,000 rat targets per sample BD-27-303 72 BD-27-302 Illumina TotalPrep RNA Amplification Kit 24 AMIL1791 Available from Ambion: 1-800-888-8804 (U.S.) 96 4393543 Includes hybridization buffers, wash buffers, and wash trays Related Products TargetAmp Nano-g Biotin-aRNA Labeling Kit Available from Epicentre Biotechnologies: 1.800.284.8474 (U.S.) TAN07924-142 Illumina, Inc. 9885 Towne Centre Drive, San Diego, CA 92121 USA 1.800.809.4566 toll-free 1.858.202.4566 tel techsupport@illumina.com illumina.com For research use only 2011 Illumina, Inc. All rights reserved. Illumina, illuminaDx, Solexa, Making Sense Out of Life, Oligator, Sentrix, GoldenGate, GoldenGate Indexing, DASL, BeadArray, Array of Arrays, Infinium, BeadXpress, VeraCode, IntelliHyb, iSelect, CSPro, GenomeStudio, Genetic Energy, HiSeq, HiScan, Eco, and MiSeq are registered trademarks or trademarks of Illumina, Inc. All other brands and names contained herein are the property of their respective owners. Pub. No. 470-2009-004 Current as of 18 January 2011
Illumina guarantees that 99.98% of the bead types will be present on any given HumanHT-12 array. This means up to five probes may be represented with only 0, 1, or 2 copies on each HumanHT-12 array. Probes were designed to cover content from NCBI RefSeq Release 38 (November 7, 2009), as well as legacy UniGene content.
AQA GCE Biology A2 Award 2411 Unit 5 DNA & Gene Expression Unit 5 Control in Cells & Organisms DNA & Gene Expression Practice Exam Questions . AQA GCE Biology A2 Award 2411 Unit 5 DNA & Gene Expression Syllabus reference . AQA GCE Biology A2 Award 2411 Unit 5 DNA & Gene Expression 1 Total 5 marks . AQA GCE Biology A2 Award 2411 Unit 5 DNA & Gene Expression 2 . AQA GCE Biology A2 Award 2411 .
Gene Expression 1. TaqMan Gene Expression Assays 2. Custom TaqMan Gene Expression Assays 3. TaqMan MicroRNA Assays 4. Use of Primer Express Software for the Design of Primer and Probe Sets for Relative Quantitation of Gene Expression 5. Design of Assays for SYBR Green I Applications Section IV.
Vector are conveniently included in the ExpiSf Expression System Starter Kit for expression of your gene of interest in ExpiSf9 cells. pFastBac 1 Expression Vector pFastBac 1 Expression Vector is a non-fusion donor plasmid that is used to clone your gene of interest using restriction enzyme digestion and ligation. Gene expression
Main purposes of this tutorial ! Provide an updated list of plant gene-expression . expression profiles ! Review considerations relevant to the use of gene expression databases ! Use web-based tools for visualization of transcriptomic data . Background ! Expression databases hosting microarray -derived data have been fundamental to study gene .
Level 2 Biology, 2013 91159 Demonstrate understanding of gene expression 9.30 am Friday 22 November 2013 Credits: Four Achievement Achievement with Merit Achievement with Excellence Demonstrate understanding of gene expression. Demonstrate in-depth understanding of gene expression. Demonstrate comprehensive understanding of gene expression.
gene expression can be regulated by modulating the degree to which the transcript is protected. 1. Initiation of transcription. Most control of gene expression is achieved by regulating the frequency of transcription initiation. 3. Passage through the nuclear membrane. Gene expression can be regulated by controlling access to or efficiency of .
3. Identify the main mechanism for turning on gene expression. Explain why control of gene expression in eukaryotic cells is like a “dimmer switch”, an “ON” switch that can be fine tuned. 4. Identify the major switch and all the fine-tuning steps that can modulate eukaryotic gene expression. 5.
The procedures in this document are for use with TaqMan Gene Expression Assays configured and preplated on TaqMan Array Plates. TaqMan Gene Expression Assays are a comprehensive collection of predesigned, preformulated primer and probe sets to perform quantitative
