TaqMan® MicroRNA Using the QuantStudio™ 12K Flex Real-Time PCR System with OpenArray® Block
OpenArray® plate technology on the QuantStudio™ 12K Flex Real-Time System helps streamline microRNA (miRNA) profiling, at a scale previously unattainable. Achieve the highest sample throughput for mid-density miRNA profiling—in a single work day, one user can process 48 samples with minimal pipetting steps and without the use of robotics.
- Broad coverage—run 3 samples per QuantStudio™ 12K Flex OpenArray® Plate translating up to 818 assays per sample (up to 758 unique assays)
- High throughput—12 samples per ~2 hr instrument run and 48 samples per 8hr working day
- Cost effective—rapid sample screening with minimal hands-on time
- Minimal sample requirement—only 200ng of total RNA needed for a full profile
Experimental Workflow: Find the Products You Need for Every Step
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Step 1: Prepare Sample
Applied Biosystems offers a variety of kits for the isolation and analysis of miRNA and other small RNAs, each is ideal for use in miRNA analysis because they enable quantitative recovery of small RNA (<200 nt) and maintenance of representative amounts of small RNA (eliminating experimental bias).
Products for Step 1: Prepare Sample
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mirVana™ miRNA Isolation Kit
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Isolate total RNA that contains small RNAs such as miRNA, siRNA, and snRNA. These small RNAs are often lost during traditional RNA isolation methods.
Step 2: Reverse Transcribe
Designed to streamline miRNA profiling, Megaplex™ RT Primers are pools of RT primers identical to those in individual TaqMan® MicroRNA Assays. These primers are ideal for global miRNA expression profiling; just two pools provide comprehensive coverage of the Sanger miRBase v14 and v15 database content for human and rodent, respectively. The content of each pool matches corresponding QuantStudio™ 12K Flex TaqMan® OpenArray® Human or Rodent MicroRNA Panels. The TaqMan® MicroRNA RT Kit provides optimal TaqMan® MicroRNA Assay performance. This kit is used to convert miRNA to cDNA prior to real-time PCR quantitation.
Products for Step 2: Reverse Transcribe
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Megaplex™ RT Primers
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Whether your profiling experiment requires ultimate sensitivity, broad coverage or both, Megaplex™ Primer Pools provide the flexibility to reach your research goals. Megaplex™ Primer Pools provide comprehensive coverage of Sanger miRBase v14 and v15 (for human and rodent, respectively), and when used with TaqMan® MicroRNA Arrays, offer an ideal miRNA profiling solution.
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TaqMan® MicroRNA Reverse Transcription Kit
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The TaqMan® MicroRNA Reverse Transcription Kit provides the necessary components for optimal performance of TaqMan® MicroRNA Assays. Components of this kit are used with the RT primer provided with the TaqMan® MicroRNA Assay to convert miRNA to cDNA.
Step 3: Amplify cDNA
Megaplex™ PreAmp Primers generate a comprehensive miRNA expression profile using as little as 1 ng of input total RNA. With content matched to specific Megaplex™ RT Primers and QuantStudio™ 12K Flex TaqMan® OpenArray® Human MicroRNA Panels, cDNA prepared by reverse transcription can be preamplified using Megaplex™ PreAmp Primers and TaqMan® PreAmp Master Mix to uniformly amplify all miRNAs present in the original sample significantly increasing assay sensitivity.
Products for Step 3: Amplify cDNA
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Megaplex™ PreAmp Primers
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For use when assay sensitivity is of the utmost importance, Megaplex™ PreAmp Primers significantly enhance the ability to detect low expressed miRNAs enabling the generation of a comprehensive expression profile using far less sample, as low as 1 ng of input total RNA.
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TaqMan® PreAmp Master Mix
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TaqMan® PreAmp Master Mix preamplifies small amounts of cDNA without introducing amplification bias to the sample. Preamplification enables you to stretch your precious sample into many more real-time PCR reactions. TaqMan® PreAmp Master Mix Kit consists of TaqMan® PreAmp Master Mix and TaqMan® Gene Expression Master Mix.
Step 4: Cycle and Image
Load sample mixes onto QuantStudio™ 12K Flex TaqMan® OpenArray® MicroRNA Panels using the QuantStudio™ 12K Flex OpenArray® AccuFill™ System. The plate is encased with an alloy bottom for easy handling. Simply apply the adhesive lid and fill with immersion fluid. Perform a real-time plate read using the QuantStudio 12K Flex Real-Time PCR System.
Products for Step 4: Cycle and Image
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QuantStudio™ 12K Flex Real-Time PCR System with OpenArray® Block
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QuantStudio™ 12K Flex Real-Time PCR System with OpenArray® Block The QuantStudio™ 12K Flex Real-time PCR System and QuantStudio™ 12K Flex OpenArray® AccuFill™ System combination can process hundreds to thousands of samples per day, enabling mid-density, high-throughput results with ease of use and without robotics.
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TaqMan® OpenArray® MicroRNA Panels
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TaqMan® OpenArray® MicroRNA Panels contain preconfigured sets of TaqMan® MicroRNA Assays for human profiling studies in the highest throughput format available from Life Technologies. The human panel lets you run 3 samples and a total of 2,274 unique TaqMan® MicroRNA Assays at once. OpenArray® plates compatible with QuantStudio™ 12K Flex Real-Time PCR system will be coming soon and available only on www.lifetechnologies.com.
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TaqMan® OpenArray® Real-Time PCR Master Mix
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TaqMan® OpenArray® Real-Time PCR Master Mix is conveniently formulated to support microRNA (miRNA) profiling using TaqMan® OpenArray® MicroRNA Panels on the QuantStudio™ 12K Flex Real-Time PCR System. OpenArray® plates compatible with QuantStudio™ 12K Flex Real-Time PCR system will be coming soon and available only on www.lifetechnologies.com.
Products for Step 5: Analyze Data
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RealTime StatMiner® Software
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RealTime StatMiner® Software from Integromics is a powerful, easy-to-use bioinformatics tool for quality control and differential expression analysis of real-time PCR data. With just a single click, the software imports raw data or Ct values from any of our real-time PCR instruments, and then guides you though every step of analysis, giving you accurate results in minutes.
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