miRNA Quantitation

Workflow Features

Ideal for quantitating individual miRNAs from a broad assortment of species, including human, mouse, rat, Drosophila, C. elegans, and Arabidopsis
Each kit includes the TaqMan® MicroRNA Assay and reverse transcription primer specific for the mature miRNA target of interest
Assays are continuously added for human, mouse, and rat species to stay aligned with the Sanger miRBase Registry

How It Works
TaqMan® MicroRNA Assays employ an innovative target-specific stem-loop reverse transcription primer to address the challenge of the short length of mature miRNA. The primer extends the 3' end of the target to produce a template that can be used in standard TaqMan® Assay-based real-time PCR. Also, the stem-loop structure in the tail of the primer confers a key advantage to these assays: specific detection of the mature, biologically active miRNA.

Products
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

mirVana™ miRNA Isolation Kit

Isolate total RNA that contains small RNAs such as miRNA, siRNA, and snRNA. These small RNAs are often lost during traditional RNA isolation methods. In addition, this kit includes reagents and instructions for the isolation of RNA that is enriched for small RNA (<200 bp), enabling more sensitive detection of low-level small RNAs.

RecoverAll™ Total Nucleic Acid Isolation Kit for FFPE

Extract total RNA or DNA from formalin- or paraformalin-fixed, paraffin-embedded (FFPE) tissues. This kit enables the molecular analysis of archived tissue samples at both the genomic and gene expression levels. With this kit, nucleic acid is obtained that is suitable for RT-qPCR, mutation screening, and microarray analyses.

TaqMan® MicroRNA Cells-to-CT™ Kit

Featuring a unique method for lysing cultured cells while removing genomic DNA and preserving RNA integrity, the TaqMan® MicroRNA Cells-to-CT™ Kit makes it possible to perform miRNA expression profiling directly from cultured cells without RNA purification.

Step 2: Reverse Transcribe RNA

Two-step TaqMan® MicroRNA Assays, containing the TaqMan® Assay and reverse transcription primer specific for the target of interest, are designed for individual miRNA targets found in a wide variety of species including human, mouse, rat, Drosophila, C. elegans, and Arabidopsis. The TaqMan® MicroRNA Reverese Transcription Kit provides optimal TaqMan® MicroRNA Assay performance. This kit is used with a single RT primer provided with each individual TaqMan® MicroRNA Assays or with Megaplex™ RT Primers to convert miRNA to cDNA prior to real-time PCR quantitation.

Products for Step 2: Reverse Transcribe RNA

TaqMan® MicroRNA Reverse Transcription Kit

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.

TaqMan® MicroRNA Assays (RT tube)

TaqMan® MicroRNA Assays quantitate microRNAs with the specificity and sensitivity of TaqMan Assay chemistry. A simple, two-step protocol requires only reverse transcription with a miRNA-specific primer, followed by real-time PCR with TaqMan probes. The assays target only mature microRNAs, not their precursors, ensuring biologically relevant results.

Step 3: Set Up Singleplex MicroRNA Assay Reaction

For the final quantitation step, the cDNA product generated in step 2 is mixed with TaqMan® Universal Master Mix II and added to the matched TaqMan® MicroRNA Assay.

Products for Step 3: Set Up Singleplex MicroRNA Assay Reaction

TaqMan® MicroRNA Assays (individual assays)

TaqMan® MicroRNA Assays quantitate miRNAs with the specificity and sensitivity of TaqMan® Assay chemistry. A simple, two-step protocol requires only reverse transcription with a miRNA-specific primer, followed by real-time PCR with TaqMan® probes. The assays target only mature miRNAs, not their precursors, ensuring biologically relevant results.

TaqMan® Universal Master Mix II

This kit contains components needed for all 5' nuclease assays using TaqMan® Assay-based PCR applications. This easy-to-use solution minimizes your assay set-up time and employs AmpliTaq Gold® DNA polymerase for enhanced performance.

Step 4: Perform Real-Time PCR Amplification

The real-time PCR can be run on any Applied Biosystems Real-Time PCR System.

Products for Step 4: Perform Real-Time PCR Amplification

StepOnePlus™ Real-Time PCR System

The Applied Biosystems® StepOnePlus™ Real-Time PCR System makes it simple and easy to get publication-ready high-quality real-time PCR results on day one. This remarkably simple 96-well, 4-color, real-time PCR system is designed with a user-friendly, yet powerful, interface for researchers of all experience levels.

7300 Real-Time PCR System

Get the Applied Biosystems advantage at an affordable price; the 7300 Real-Time PCR System balances exceptional quality with budget sensitivity.

7500 Real-Time PCR System

The Applied Biosystems® 7500 Real-Time PCR System is a powerful platform for labs requiring superior performance and maximum dye versatility. This third-generation platform features an innovative optical system that enhances sensitivity and lets you access a broader range of fluorophores. The variable excitation capability allows greater sensitivity for longer wavelength (red) dyes.

7500 Fast Real-Time PCR System

The Applied Biosystems® 7500 Fast Real-Time PCR System offers maximum performance in the minimum time. Fully optimized for Fast cycling, the 7500 Fast System delivers high-quality results in as little as 30 minutes.

7900HT Fast Real-Time PCR System

The 7900HT Fast Real-Time PCR System is the only real-time quantitative PCR system that combines 384-well plate compatibility with fully automated robotic loading — and also offers optional Fast real-time PCR capability. Acknowledged as the gold standard in real-time PCR, the 7900HT System combined with TaqMan® Genomic Assays enables you to achieve unprecedented throughput and flexibility, allowing you to pursue projects beyond the scope of previous real-time instruments.

Step 5: Analyze Data

Integromics RealTime StatMiner® Software is a powerful, easy-to-use bioinformatics tool for quality control and differential expression analysis of real-time PCR data. The software easily imports raw data or Ct values from any Applied Biosystems® Real-Time PCR System and guides you though every step of analysis for accurate results in minutes.

Products for Step 5: Analyze Data

RealTime StatMiner® Software (optional)

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 through every step of analysis, giving you accurate results in minutes.
Literature
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- Leading the Way in MicroRNA
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