SNP基因型分析

Powerful, Proven Chemistry Whether your genotyping studies require targeted detection of essentialSNPs, or the flexibility for choosingSNPs for mapping, TaqMan SNPGenotyping Assays are the technologyof choice. Proven TaqMan probes, whichincorporate minor groove binder(MGB) technology at the 3’ end,deliver superior allelic discrimination. The MGB molecule binds to the minor groove of the DNA helix, improvinghybridization-based assays by stabilizing the MGB-probe/template complex. The increased binding stabilization permits the use of probes as short as 13 basesTaqMan ®SNP Genotyping AssaysTaqMan ® SNP Genotyping Assays from Applied Biosystems provide a highly flexible technology for detection of poly-morphisms within any genome. With the simplest workflow available, TaqMan ® Assays are the quickest way to generate genotyping data. Based on powerful TaqMan ® probe and primer chemistry and designs, and coupled to dependable Applied Biosystems instruments and software, these Made-to-Order assays produce high-confidence results. These TaqMan Assays are ideal for genotyping applications including screening, associa-tion, candidate region, candidate gene,or fine-mapping studies.Content-rich marker-selection toolssimplify study design and help youselect from a library of human andmouse assays. This library includesover 4.5 million genome-wide humanassays (of which 3.5 million areHapMap SNP-based assays, 160,000 are validated assays, and over 70,000 are coding region assays) and 10,000 mouse assays. We also offer over 2,600 Inventoried Drug MetabolismGenotyping Assays. Additionally, Custom TaqMan ® SNP Genotyping Assays let you create your own confidential assays by submitting target SNP sequences for any genome. Let TaqMan SNP Genotyping Assays accelerate the pace of yourdiscovery by eliminating time-consuming experimental design and optimization.Figure 1. Allelic discrimination is achieved by the selective annealing of TaqMan MGB probes.Visit for more information.Over 10,000 mouse SnP Genotyping assaysThe Mouse TaqMan ® Pre-Designed SNP Genotyping Assays collection consists of over 10,000 assays, and can also be supplemented with assays designed using our Custom TaqMan ® SNP Genotyping Assays Service.T aqman ® Drug metabolism Genotyping assays Collection Over 2,600 assays that target high value polymorphisms in over 220 drug metabolism genes. These assays have proven performance in four different ethnic populations, comprised of 45 individuals each. To enable easyidentification, these assays have been mapped to the common public allele nomenclature Web sites where possible. Visit for more information.for improved mismatch discrimination and greater flexibility when designing assays for difficult or variable sequences. In addition to SNP detection, TaqMan probes can be designed to detect multiple nucleotide polymorphisms (MNPs) and insertion/deletions (InDels).Detection is achieved with proven 5’ nuc- lease chemistry by means of exonuclease cleavage of a 5’ allele-specific dye label, which generates the permanent assay signal (Figure 1). All MGB probes include a nonfluorescent quencher (NFQ) that virtually eliminates the backgroundfluorescence associated with traditional quenchers and provides a greater signal- to-noise ratio for superior assay sensitivity.T aqman ® SnP Genotyping assays CollectionTaqMan SNP Genotyping Assays are the world’s largest collection of single-tube, ready-to-use SNP assays available.The TaqMan SNP Genotyping Assays library consists of two human and one mouse assay collections, and can be supplemented with assays designed using our Custom TaqMan ® SNP Genotyping Assays Service (Table 1).Over 4.5 million Human SnP Genotyping assaysThis assay group contains over 4.5 million genome-wide SNPs providing unprecedented marker coverage.Included in this collection are 160,000 validated assays that have roughly 10-kb spacing across gene regions.These assays were subjected to a minor allele frequency test in 2–4 ethnic populations (45 individual samples per ethnic group) and offer the highest success rate because of this extensive testing. Over 70,000 assays are also included for the detection of nonsyn-onymous SNPs in coding regions, includ-ing many putative functional SNPs.Human assays Non-human Number of Reactions at Reactions at Assay mix Availabilitypart numbers assays part SNPs 5 μL volume 25 μL volume formulationnumbers (384-well plate) (96-well plate)TaqMan ® Pre-Designed SNP Genotyping Assays for Human and Mouse Small-scale 4351379 4351384* >4,500,000 1,500 300 40X Made-to-Order Medium-scale 4351376 4351382* >4,500,000 5,000 1,000 40X Made-to-Order Large-scale43513744351380*>4,500,000 12,0002,40080XMade-to-OrderTaqMan ® Drug Metabolism SNP Genotyping Assays Small-scale4362691N/A >2,600 750 150 20X InventoriedCustom TaqMan ® SNP Genotyping Assays Small-scale 4331349 4332077 ∞ 1,500 300 40X Made-to-Order Medium-scale 4332072 4332075 ∞ 5,000 1,000 40X Made-to-Order Large-scale43320734332076∞12,0002,40080XMade-to-OrderAll assays are quality control tested using a mass spectrophotometer to verify sequence and yield. All assays have one VIC ® and one FAM ™ dye-labeled probe and two target-specific primers. All assays, except Custom Assays, undergo bioinformatics evaluation of target SNP sequences.Functional testing against 20 unique genomic DNAs is performed on all human SNP Genotyping Assays. Validation testing against three populations with ~45 samples/population was performed on all Validated and Drug Metabolism Assays.All Validated and Coding Assays that were previously held in inventory are now available as Made-to-Order Pre-Designed Assays. Their assay IDs remain unchanged.*Over 10,000 mouse assays available.All TaqMan SNP Genotyping Assays were generated using next-generation algorithms from the Applied Biosystems bioinformatics pipeline. Bioinformatics evaluation of target SNP sequences includes the masking of adjacent SNPs and ambiguous bases so that assay design and subsequent performanceis not affected by underlying sequence quality. Lastly, the assay designs are aligned to the human genome using BLAST to ensure the assay is uniquely binding and targets only the intended polymorphism. As the Custom TaqMan SNP Genotyping Assay Service is a confidential and secure service, custom-ers perform their own bioinformatics analysis prior to submission of sequence for assay design.Free marker Selection T ools SNPbrowser™ Software for Human SNPs SNPbrowser™ Software for human SNPs simplifies study design by facilitating easy and intuitive selection of the optimal SNP assay set for each project. Y ou can download free SNPbrowser Software at /snpbrowser SNPbrowser Software provides a physical map view of the human genome.Two different genotype data sources are incorporated into the software togenerate unique linkage disequilibrium (LD) maps and haplotype block informa-tion. These two sources are the public HapMap Project and Applied Biosystems’ 20+ million genotypes generated during the development of the 160,000 TaqMan Validated SNP Genotyping Assays.This genotyping data also facilitates the use of tagging SNP methods, which allow more affordable study design by reducing the number of SNPs while conserving study statistical power (Figure 2). SNPbrowser Software allows you to optimize study design based on LD patterns, minor allele frequency require-ments, total number of cases and controls, and statistical power. Finally, the software includes a SNP density tool that prioritizes the selection of validated SNPs while supplementing with additional SNPs to fulfill the markerdensity requirements for your study.When you have identified your SNPsof interest, order TaqMan SNPGenotyping Assays by directly uploadingAssay IDs from the software to theApplied Biosystems Web site by goingto Mouse SNPbrowser™ SoftwareComplimentary Mouse SNPbrowser™Software enables efficient and easyselection of appropriate SNP setsto discriminate between strains ofinterest, with specific applications ingenotype/phenotype mapping and strainverification. It visualizes informativeSNPs that distinguish two selectedmouse strains at a user-definableresolution. There are ~10,000 mouseSNPs genotyped in 44 of the mostcommon strains.Mouse SNPbrowser selects fromPre-Designed SNP Genotyping assaysusing either TaqMan® Assay or SNPlex™Genotyping System Assay collectionsavailable from the Applied Biosystemsonline store. Y ou can download freeMouse SNPbrowser Software from/mousesnpbrowserMouse SNPbrowser Software supportsgenetic monitoring, genetic mapping,and speed congenics applications byoffering a variety of tools to search, view,export SNP information, and purchaseassays. The software displays the mousechromosome map so users can viewSNP location and obtain National Centerfor Biotechnology Information (NCBI)data for SNPs of interest. The softwareallows SNP selection for two applications:genetic monitoring and genetic mapping. Figure 2. Identifying Optimal Subsets of SNPs with the SNP Wizard. (A) Search and visualize your gene of interest, then activate the SNP Wizard. (B) Choose a tagging SNP method and set parameters to reducethe number of SNPs needed to represent common haplotypes. A summary panel appears that displays algorithm results and allows you to easily adjust your parameter selection. Selected tagging SNPs are shownin red on the graphical map.A.B.For genetic monitoring, the software selects optimal SNPs to distinguish between any number of strains. SNPs can be selected for each chromosome or across the genome. For genetic mapping, the software segments each chromosome into evenly sized blocks based on user-specified mapping resolu-tion, and searches for distinguishing SNP in each block to generate evenly-spaced SNP coverage throughout the genome. Mouse SNPbrowser Software also features a SNP wizard to assist defining search criteria. A help text file is provided within the tool to assist in familiarizing users of the extensive functionality offered.Custom assay Service for any Possible SnPCustom TaqMan SNP Genotyping Assays are available for any possible SNP in any organism. This service can generate assays for the detection of SNPs, MNPs, or insertions/deletions of up to six bases. Custom TaqMan SNP Genotyping Assays provide customers with a complete service that includes secure and confidential ordering, assay design and manufacturing, quality-control testingfor synthesis accuracy and formulation completeness. Additionally, custom human assays are subjected to a func-tional test on 20 unique DNA samples. Our complimentary File Builder Software makes it easy to submit your targetsfor design through our custom assay service. Download the latest versionof File Builder Software at/filebuilder Quality Design and manufacturing Probes and primers used in TaqMan SNP Genotyping Assays are designed byour rigorous bioinformatics pipeline. This proprietary group of algorithms has generated millions of TaqMan Assay designs by utilizing heuristic design rules deduced from both manufacturing and assay performance data. All assays are designed to perform under universal reaction conditions, as calculated probe and primer melting temperaturesare consistent and include contributionsfrom associated probe conjugates(i.e., dyes, MGB).After manufacturing, assay componentsundergo extensive laboratory testing atour state-of-the-art manufacturing facility(Figure 3). Quality-control testing includesmass spectrometry for sequenceFigure 4.A simple workflow and reliable instruments combine to generate fast, high-confidence results. Figure 3. Applied Biosystems high-throughput manufacturing facility.verification and formulation assessments of probe and primer concentrations. Additionally, we functionally test all human SNP genotyping assays with an allelic discrimination test.Simple Workflow for Quick Results TaqMan SNP Genotyping Assaysconstitute the simplest SNP genotyping technology available. Applied Biosystems delivers your ready-to-use SNP geno-typing assay in a convenient, single-tube format. The rest is easy. Just combine the assay with TaqMan ® Genotyping Master Mix or TaqMan ® Universal PCR Master Mix and your purified DNA sample. There is no need to optimize temperature or concentrations of probes, primers, or salts, because all assays run under universal reagent concentrations and thermal cycling conditions.After thermal cycling, no transfers, wash-es, or addition of reagents is required, and the plate remains sealed; just read the plate and analyze the genotypes (Figure 5). This reduces the chance of contamination, sample mix-up, and sample loss. Because of the simplicity in chemistry, you can easily automate the reaction for massively parallel genotyping studies, readily increasing the number of assays, number of samples, or both. Additionally, the analysis software allows you to auto-call genotypes, minimizing manual intervention.Reliable Real-Time PCR Platforms Applied Biosystems offers a suite of superior instrument platforms for processing and analyzing TaqMan SNP Genotyping Assays. These instruments, which meet all throughput needs and budgets, include the GeneAmp ® PCR System 9700 and Veriti ™ Thermal Cyclers and the Applied Biosystems 7300, 7500, 7500 Fast, 7900HT Fast, StepOne ™ or StepOnePlus ™ Real-Time PCR Systems. Following PCR amplification, an endpoint read can be performed on any Applied Biosystems Real-Time PCR System. All of these dependable instruments offer the advanced multicolor detection capabilities required for highly accurate and reproduc-ible allelic discrimination assays.Figure 5.The SNP auto-caller feature automatically determines sample genotypes and displays data.Figure 6. The flexible, high-throughput Applied Biosystems 7900HT Fast Real-Time PCR System.Table 2. InSTRumenT CaPaCITyInstrument PlatformsCapacityApplied Biosystems GeneAmp ® 60-, 96-, Dual 96-, or Dual 384-well PCR System 9700 Thermal Cycler blocksApplied Biosystems Veriti ™ Thermal Cycler 60-, 96- (Standard or Fast),or 384-well block Applied Biosystems 7300 Real-Time PCR System 96-well blockApplied Biosystems 7500/7500 Fast 96-well block (Standard and Fast) Real-Time PCR SystemApplied Biosystems 7900HT Fast 384-well block (Standard and Fast) Real-Time PCR System Applied Biosystems StepOne ™ 48-well block (Standard and Fast) Real-Time PCR SystemApplied Biosystems StepOnePlus ™ 96-well block (Standard and Fast)Real-Time PCR SystemHeadquarters850 Lincoln Centre Drive | Foster City, CA 94404 USA Phone 650.638.5800 | Toll Free 800.345.5224 International SalesFor our office locations please call the division headquarters or refer to our Web site at/about/offices.cfmFor Research Use Only. Not for use in diagnostic procedures. NOTICE TO PURCHASER: LIMITED LICENSEA 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.TaqMan SNP Genotyping Assays contain Licensed Probes. Use of these products is covered by one or more of the following US patents and corresponding patent claims outside the US: 5,538,848, 5,723,591, 5,876,930, 6,030,787, 6,258,569, and 5,804,375 (claims 1-12 only). The purchase of these products includes a limited, non-transferable immunity from suit under the foregoing patent claims for using only the amount of product specified for the purchaser’s own internal research. Separate purchase of an Authorized 5’ Nuclease Core Kit would convey rights under the applicable claims of US Patents Nos. 5,210,015 and 5,487,972, and corresponding patent claims outside the United States, which claim 5’ nuclease methods. No right under any other patent claim and no right to perform commercial services of any kind, including without limitation reporting the results of purchaser’s activities for a fee or other commercial consideration, is conveyed expressly, by implication, or by estoppel. These products are 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.© 2008. Applied Biosystems. All Rights Reserved. Information subject to change without notice. Applera, Applied Biosystems, AB (Design), GeneAmp, and VIC are registered trademarks and FAM, StepOne, StepOnePlus, and Veriti are trademarks of Applera Corporation in the US and/or certain other countries.AmpliTaq Gold and TaqMan are registered trademarks of Roche Molecular Systems, Inc.Printed in the USA. 06/2008 Publication 135PB03-02Data analysis SoftwareThe sophisticated software package provided with all Applied Biosystems’ Real-Time PCR Systems facilitates experimental set-up, data collection, and assay performance analysis. The SDS software uses an advanced multi-component algorithm to calculate distinct allele/marker signal contributions from fluorescence measurements for each sample well during the assay plate read. The SNP auto-caller feature automatically determines samplegenotypes and generates a cluster plotFigure 7. The Applied Biosystems Web site ( ) facilitates convenient online ordering and multiple search options for all our genotyping assays, including keyword, batch, and location searches.that helps you visualize data across samples or populations (Figure 5).For large-scale studies, you can cycle samples on multiple Dual 384-Well GeneAmp ® 9700 or Veriti ™ Thermal Cyclers and then analyze them on a 7900HT System. The 7900HT System (Figure 6) facilitates high-throughput applications by allowing large sample batches (up to eighty-four 384-well plates) to be processed withoutmanual intervention using the optional Automation Accessory. Following data collection, you can accelerate andsimplify high volume data transfer and analysis with the optional SDS version 2.2 Enterprise Edition Software Suite.Ordering InformationSelecting and ordering TaqMan SNP Genotyping Assays is as simple as “point and click.” Use SNPbrowserSoftware to select the most informative SNPs for your genotyping studies. As you identify SNPs of interest, simply upload your selected TaqMan SNP Genotyping Assays to the Applied Biosystems Web site for ordering.The Applied Biosystems Web site (Figure 7) enables you to search for, select, and order from our catalog of over 4.5 million TaqMan SNP Genotyping Assays. Y ou can search for SNPs using several criteria options: National Center for Biotechnology Information (NCBI) gene ID, NCBI SNP reference ID (rs#) and gene symbol. Y ou can further refine your search by using SNP type (i.e., intragenic, 5’ or 3’ UTR, chromosome, etc.).Our Custom TaqMan SNP Genotyping Assays Service is provided to supply you with any SNP not available from our catalog, including any non-humanorganism. This service designs assays for all possible SNP , MNP , and InDeltargets. Our complementary File Builder Software conveniently formats your target sequence for submission to our manufacturing facilities. To order any custom assays, simply upload yoursubmission file to our Web site, or emailthe file to your local sales office.。