RealTime ready Custom RT-qPCR Assays and Panels for any Human, Mouse, or Rat Gene

Transcription

1 RealTime ready Custom RT-qPCR Assays and Panels for any Human, Mouse, or Rat Gene

2 RealTime ready qpcr Assays and Panels 4 RealTime Ready Configurator 7 Search by Focus Panel 8 Search by Pathway 10 Ordering Information 11 For life science research only. Not for use in diagnostic procedures. License Disclaimer: A license to perform the patented 5 Nuclease Process for research is obtained by the purchase of (i) both Licensed Probe and Authorized 5 Nuclease Core Kit, (ii) a Licensed 5 Nuclease Kit, or (iii) license rights from Applied Biosystems. This product contains Licensed Probe. Its purchase price includes a limited, non-transferable immunity from suit under U.S. Patent No. and 5,804,375 (claims 1-12 only (exp. 9/8/2015)) and corresponding patent claims outside the United States. The purchase of this product includes a limited, non-transferable immunity from suit under the foregoing patent claims for using only this amount of product for the purchaser s own internal research. The right to use this product in the 5 Nuclease Process under the corresponding patent claims outside the United States, can be obtained through purchase of an Authorized 5 Nuclease Core Kit. Except under separate license rights available from Applied Biosystems, no right under any other patent claim, or to perform commercial services of any kind, including without limitation reporting the results of purchaser s activities for a fee or other commercial consideration, or to sublicense, repackage with other products, or resell in any form, is conveyed expressly, by implication, or by estoppel. This product is for research use only. Diagnostic uses under Roche patents require a separate license from Roche. Further information on purchasing licenses may be obtained from the Director of Licensing, Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404, USA. ProbeLibrary is covered by US and other patent applications owned by Exiqon A/S. Locked Nucleic Acids (LNA) are covered by U.S. Patents No US 6,794,499 (exp. 9/11/2018), US 6,670,461 (exp. 9/11/2018), US 6,268,490 (exp. 3/9/2018) & US 6,770,748 (exp. 3/9/2018) and other patents and patent applications owned by Exiqon A/S and Prof. Takeshi Imanishi. The quenche Parts of the Software used for the LightCycler 480 System are licensed from Idaho Technology Inc., Salt Lake City, UT, USA. This product is covered by one or more of U.S. 6,197,520 (exp. 8/13/2019), 6,303,305 (exp. 3/30/2019), 6,387,621 (exp. 9/8/2019), 6,503,720 (exp. 2/20/2021), 6,730,501 (exp. 3/13/2022) and corresponding claims in their non-u.s. counterparts, owned by Roche Diagnostics GmbH and/or licensed from Idaho Technology, Inc. This LightCycler 480 Real-Time PCR System is a real-time thermal cycler licensed for research use only under U.S. Patent No. 6,814,934 (exp. 5/2/2011) and corresponding claims in its non-u.s. counterparts, and under one or more of U.S. Patents Nos. 5,475,610 (exp. 12/12/2013), 5,602,756 (exp. 3/14/2011), 6,703,236 (exp. 11/29/2010), 7,238,517 (exp. 11/29/2010), 7,504,241 (exp. 11/29/2010), 7,537,377 (exp. 2/9/2020) or corresponding claims in their non-u.s. counterparts, owned by Applied Biosystems, LLC. No rights are conveyed expressly, by implication or estoppels to any other patent claims. The LightCycler 480 Real-Time PCR System is not intended for in vitro diagnostic use in the U.S. For further information on purchasing additional rights, contact the Director of Licensing at Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California, 94404, USA. No right is conveyed expressly, by implication, or by estoppels under any other patent claim, such as claims to apparatus, reagents, kits, or methods such as 5 nuclease methods. License rights to practice PCR methods under the foregoing Roche patents for research and other non-in vitro diagnostic applications may be purchased from Applied Biosystems or may be obtained by purchasing licensed reagents from Roche, Applied Biosystems, or other Authorized third party. For information on purchasing licenses for research and other non-in vitro diagnostic applications, contact the Director of Licensing at Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404, USA. A license to practice PCR methods with real-time detection under patents of F. Hoffmann- La Roche Ltd, and Roche Molecular Systems ( Roche ) for in vitro diagnostic applications may be purchased from Roche or may be obtained by purchasing in vitro diagnostic reagents from Roche or any other Authorized third party. The LightCycler 480 Real-Time PCR System is not intended for in vitro diagnostic use in the U.S.This product is covered in-part by US 5,871,908 (exp. 12/16/2014) or any foreign equivalents, co-exclusively licensed from Evotec OAI AG. The purchase price includes a license to practice the methods covered by US 5,871,908 (exp. 12/16/2014) by using the product. Purchase of this product, however, does not convey to the purchaser a license or right to (i) commercially make, have made or sell reagents and/or kits, or (ii) buy or use reagents and/or kits provided by a third party used in conjunction with the product or any other thermocycler to practice the methods covered by US 5,871,908 (exp. 12/16/2014) or any foreign equivalents. 2

3 RealTime ready Complex Gene Expression Analysis of Human, Mouse, and Rat Genes, Made Simple Maximize efficiency Select assays for your targets of choice from gene lists of functionally related gene families or biological pathways. Order them as single assays or configure your own custom panel on a LightCycler 480 Multiwell Plate. Ensure optimal results Rely on function tested qpcr assays based on proven Universal ProbeLibrary technology, with detailed bioinformatic background information and novel error-detection features. Get the complete picture Discover the full functionality of your targets in complex biological pathways, and reveal their influence on processes leading to diseases or disorders. 3

4 RealTime ready qpcr Assays and Panels Get a Head Start on Your Gene Expression Analysis RealTime ready RT-qPCR Assays for any human, mouse, or rate gene of your choice are based on highly specific, LNA-substituted short hydrolysis probes from the Universal ProbeLibrary (UPL). Rigorously tested on the LightCycler 480 Instrument with species-specific universal biological samples, each assay is experimentally qualified to perform according to strict criteria derived from the MIQE* guidelines. Linear dynamic range of at least 3 logs PCR efficiency values of 90% to 110% R² value of standard curve between 0.99 and 1.00 High amplification specificity single band in agarose gel analysis RealTime ready Assays, like all UPL assays, can be used successfully on any real-time PCR system under standard PCR conditions. Linearity plot (Cp vs log concentration) over five 10-fold dilution steps for 12 assays from the Human Apoptosis Pathway. Gel analysis of 48 RealTime ready Assays of the Human GPCR gene family. Dilution series of 50 ng to 5 pg of the Human RAN/RANPI gene, analyzed in triplicates. * Minimum Information for Publication of Quantitative Real-Time PCR Experiments 4

5 Assay Design Characteristics RealTime ready Assays are designed using the proprietary ProbeFinder Software, which is based on optimized and verified algorithms. For a particular gene, one representative transcript is chosen for assay design. The selection of this transcript is based on annotation and cross-references within two of the main public gene/transcript annotation resources, Ensembl and Entrez Gene. This procedure ensures comprehensive coverage of most known splice variants of a certain gene. Most RealTime ready Assays are intron-spanning, enabling reliable quantification unbiased by genomic DNA residues in the sample material. Assay Details For each RealTime ready Assay, the following information is provided: Accession ID of the transcript for which the assay was designed Location of the assay on the selected transcript, as well as on all other splice variants that were mapped to the assay by in silico prediction Amplicon length Intron/exon boundaries for intron-spanning assays Primer sequence and UPL probe number (provided upon assay delivery) Symbol Assay ID Chromosomal position Main transcript used for design Start/end in main transcript Intron Exon 3'UTR Intron-spanning 5

6 RealTime ready Custom Panels for the LightCycler 480 System Custom panels on LightCycler 480 Multiwell Plates 96 or 384 are composed of RealTime ready Assays of your choice. A multitude of different layouts (see Figures 1 and 2) is available for configuration. Assays are pre-plated and dried down for a reaction volume of 20 µl on 96-well plates and 10 µl on 384-well plates; simply add sample cdna and LightCycler 480 Probes Master for fast and reliable results. On each panel, up to three wells are reserved for reference gene assays of your choice, matching the organism you have selected for your assays. In addition, most panel layouts feature pre-plated error detection for the respective organism. Figure 1: Example RealTime ready Custom Panel Visit to view a short video demonstration and access detailed information on how to easily customize your panels. In addition, you can easily start browsing the Configurator at Figure 2: Example RealTime ready Custom Panel Error Detection Feature 1 Reference Genes are mandatory controls on each custom panel that need to be selected during the custom panel configuration process. 2 RT-Positive Control checks for quality of initial RNA and the efficiency of the RT step. It consists of three assays, each targeting different positions of the same transcript: 3 end, center region, and 5 end. 3 RT-Minus Control detects residual genomic DNA. It consist of two identical assays in two wells: the first well is for the cdna sample and the second well is for the RNA sample

7 RealTime ready Configurator The RealTime ready Configurator online design tool features an intuitive graphical interface and a large variety of layouts (with or without controls) that can be filled in with assays of your choice. Your configuration progress is automatically stored in your passwordprotected account at every step; your panel content will be available for download after your purchase is completed. Browse the Configurator at Search Functions Assay selection and configuration of custom panels is easy with the comprehensive search functions of the RealTime ready Configurator. Search by Pathway: Select assays for your targets from interactive maps of cellular or signaling pathways. Search by Focus Panel: Select assays for your targets from preselected lists of large gene families or functionally related groups. Search by Keyword: Search by gene name, gene symbol, public accession IDs, biological process, protein function, or a combination of up to 5 keywords. Use Batch Upload to search for up to 384 different accession IDs. 7

8 Search by Focus Panel RealTime ready Focus Panels The RealTime ready Configurator features Focus Panels with pre-selected assays for genes from a variety of pathways, functional groups, and gene superfamilies. The table below shows a selection of Focus Panels, grouped into different research topics and biological pathways. Cell Death Cell Cycle Development and Differentiation Drugable Genome Angiogenesis Cell Cycle Checkpoints Angiogenesis ABC Transporter Apoptosis Cyclin E Lifecycle Inhibition of Angiogenesis GPCR Signaling Caspase Activation G1/S Transition Apoptosis Ion Channels - Ligand Gated Execution of Apoptosis M-Phase Cell Cycle Ion Channels - Voltage Gated Induction of Apoptosis Protein Degradation during Cell DNA Damage and Repair JNK Signaling Cycle Negative Regulation of Apoptosis S-Phase EGF Signaling Nuclear Receptors Positive Regulation of Apoptosis Mitotic Cell Cycle Downregulation of EGF Signaling Proteases Apoptosis - Selection 384 Cell Cycle - Selection 96 Translation Initiation Protein Kinases Apoptosis - Selection 96 Apoptosis GPCRs Protein Kinases - AGC Group Proteolytic Processes During Chromatin Modification Innate Immunity Protein Kinases - Atypical Group Apoptotic Cell Death DNA Cleavage During Apoptotic Eukaryotic Translation Initiation Lipases/Esterases Protein Kinases - CAMK Group Cell Death Cell Cycle Innate Immunity PDGF Signaling Protein Kinases - CK1 Group Cytochrome P450 Structures Integrin Family Proteases Protein Kinases - CMGC Group Hemostasis Neurotrophin Signaling Protein Kinases Protein Kinases - Other Group Integrin Family Nuclear Receptors Stem Cell Research Protein Kinases - RGC Group PDGF Signaling Proteases Generation of IPSCs Protein Kinases - STE Group Proteases Protein Kinases Hematopoietic Lineage Marker Protein Kinases - TK Group Genes Transcription Factors Ectoderm Marker Genes Protein Kinases - TKL Group Endoderm Marker Genes Transcription Factors Mesoderm Marker Genes Cardio Panel Small GTPase Mediated Signal Transduction Steroid Hormone Receptor Signaling Pathway TNFR Transcription Factors Tyrosine Kinase Receptor Signaling VEGF Signaling Cardiac Cell Characterization Smooth Muscle Marker 8

9 Use the Configurator to browse the contents of each of the gene lists below, and use the Search by Focus Panel function to create your own custom panel with function tested assays. You can either load a complete gene list to your custom panel or create your own selection by picking the targets from one or more Focus Panels. Immune Response Oncology Signal Transduction Apoptosis Angiogenesis ABC Transporter FLT3 Signaling Apoptosis Angiogenesis GPCR Signaling Cell Cycle Apoptosis Hemostasis Cytochrome P450 Structures EGF Signaling Inhibition of Angiogenesis DNA Damage and Repair Downregulation of EGF Signaling Immune System EGF Signaling FLT3 Signaling Innate Immunity Innate Immunity GPCR Signaling Integrins Integrin Family Interleukin Signaling Interleukin Family Ion Channels JNK Signaling Interleukin Receptors Interleukin Signaling Neurotrophin Signaling JNK Signaling Neurotrophin Signaling PDGF Signaling NF-KB Signaling Nuclear Receptors RHO GTPase Signaling NF-KB Target Genes NF-KB Signaling SH2 Domain Proteins Signaling in Immune System PDGF Signaling Signaling in Immune System SH2 Domain Proteins Proteases Small GTPase Mediated Signal Transduction Hematopoietic Lineage Marker Genes Protein Kinases Steroid Hormone Receptor Signaling Pathway TNFR RHO GTPase Signaling Translation Initiation Translation Initiation Signaling in Immune System Tyrosine Kinase Receptor Signaling Small GTPase Mediated Signal VEGF Signaling Transduction Steroid Hormone Receptor WNT Signaling Signaling Pathway Stem Cell Research Transcription Factors VEGF Signaling WNT Signaling 9

10 Search by Pathway The Search by Pathway function allows you to select your targets from a variety of interactive maps of cellular or signaling pathways. These maps immediately reveal possible interactions of your selected targets with other genes. Simply click on the genes of interest to add them to your assay search. 10

11 Ordering Information Product Cat. No. Pack Size RealTime ready Custom Assays RealTime ready Custom Panels RealTime ready Focus Panels RealTime ready Human Reference Gene Panel, 96 RealTime ready Human Reference Gene Panel, 384 RealTime ready Human Apoptosis Panel, 96 RealTime ready Human Apoptosis Panel, 384 RealTime ready Human ABC Transporter Panel, 96 RealTime ready Human ABC Transporter Panel, 384 RealTime ready Human Cell Cycle Regulation Panel, 96 RealTime ready Human Cell Cycle Regulation Panel, 384 Visit: Visit: plates (each containing 96 assays) plates (each containing 384 assays) plates (each containing 96 assays) plates (each containing 384 assays) plates (each containing 96 assays) plates (each containing 384 assays) plates (each containing 96 assays) plates (each containing 384 assays) RealTime ready Human GPCR Panel, plates (each containing 96 assays) RealTime ready Human GPCR Panel, plates (each containing 384 assays) RealTime ready Human Nuclear Receptor Panel, 96 RealTime ready Human Nuclear Receptor Panel, 384 Real-Time PCR Reaction Mixes plates (each containing 96 assays) plates (each containing 384 assays) RealTime ready DNA Probes Master x 1 ml (5x conc.) LightCycler 480 Probes Master x 1 ml (2x conc.) Additional Products RealTime ready Cell Lysis Kit Kit (500 reactions) Transcriptor First Strand cdna Synthesis Kit (50 reactions) Kit Transcriptor Universal cdna Master reactions LightCycler 480 Sealing Foil foils Roche Applied Science offers a large selection of reagents and systems for life science research. For a complete overview of related products and manuals, visit and bookmark our homepage, and our Special Interest Sites including: RealTime ready Custom Assays and Panels: The Universal ProbeLibrary: The LightCycler System family for real-time PCR: Amplification Innovative tools for PCR: Trademarks: LIGHTCYCLER, REALTIME READY, and NIMBLEGEN are trademarks of Roche. Exiqon, LNA ProbeFinder and ProbeLibrary are registered trademarks of Exiqon A/S, Vedbaek, Denmark. For life science research only. Not for use in diagnostic procedures. 11

12 Published by Roche Diagnostics GmbH Sandhofer Straße Mannheim Germany 2011 Roche Diagnostics. All rights reserved