PCR and qPCR
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Strategies for optimizing high throughput qPCR for expression profiling

Real-time, reverse transcription-quantitative PCR (RT-qPCR) is the gold standard for gene expression analysis. This method allows researchers to generate large amounts of data from small sample sizes in just a few hours. These same characteristics should make it an ideal technique for evaluating a large number of samples for hundreds of expression markers. However, careful attention to experimental design and data analysis are needed to ensure that you obtain accurate and precise results from high throughput experiments. These precautions can also save you time and money.

Whether you are new to or experienced with high throughput gene expression profiling, listen to a joint webinar from IDT and the TATAA Biocenter to get valuable tips for designing and analyzing experiments from a prominent qPCR expert. Dr. Mikael Kubista, founder of the TATAA Biocenter and coauthor of the MIQE guidelines [1], shares his knowledge about the major challenges of high throughput expression profiling. Topics include optimizing and validating assays, sample quality testing, merging multi-plate measurements into a common analysis, as well as measuring and compensating for background due to genomic DNA. Along the way, you will learn about advantageous QC products from TATAA Biocenter and robust qPCR assays and controls from IDT.


Reference


Speaker profile

mikael kubista

Dr Mikael Kubista is the founder and CEO of the TATAA Biocenter (www.tataa.com) and head of the Department of Gene Expression at the Institute of Biotechnology of the Czech Academy of Sciences. He was a pioneer in the development of quantitative real-time PCR (qPCR) and introduced qPCR for single-cell expression profiling. Dr Kubista coauthored the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines [1] and is a member of the CEN and ISO workgroups that are drafting the forthcoming technical specifications and guidelines on the preanalytical processes in molecular diagnostics. At TATAA, he has led the development of high throughput expression profiling and pioneered multiway profiling. Dr Kubista has also developed qPCR tomography for intracellular expression profiling and was in the TATAA team that pioneered multianalyte single-cell profiling.

Product focus

Assays and controls for qPCR from IDT

PrimeTime® qPCR Assays
  • 5′ nuclease, probe-based assays—the gold standard for quantitative gene expression studies
  • Primer-based assays—designed for intercalating dye experiments

Create custom assays that are designed using our proprietary bioinformatics algorithms for any target and to your specific parameters. Alternatively, select one of our designed assays for human, mouse, and rat mRNA targets that are supported by our bioinformatics algorithms and up-to-date sequence information.

Learn more at www.idtdna.com/PrimeTime.

gBlocks® Gene Fragments

gBlocks Gene Fragments are double-stranded, 125–2000 bp DNA molecules. They are ideal for use as qPCR controls and standards, as well as for gene construction and editing applications. These affordable gene fragments are sequence-verified, ship in a few working days, and save laboratory time.

Learn more at www.idtdna.com/gBlocks.


High throughput qPCR kits and reagents from TATAA Biocenter

Visit www.tataa.com/products to learn more about kits, reagents, and controls, such as the ValidPrime® Kit as an alternative to –RT controls or the Interplate calibrator kit.

Additional reading

Selecting qPCR assays in plates for high throughput and large-scale studies—Use these tricks for easy ordering of qPCR assays in plates.

Multiplex qPCR—how to get started—Learn how multiplex qPCR can save sample, reagent cost, and time. The article provides recommendations for multiplex qPCR assay design and experimental setup.

Increase sensitivity and precision in your qPCR experiments—Use double-quenched probes to decrease background, and increase sensitivity and precision in your qPCR experiments.

Improved pathogen detection by multiplex RT-qPCR—Citation summary: See how gBlocks® Gene Fragments can help optimize multiplex qPCR for pathogen detection in human clinical samples.

Browse our complete list of DECODED newsletter articles on PCR applications.

Visit the DECODED Online newsletter for articles on molecular biology and genomics applications.

Author: Maureen Young, PhD, is a scientific writer at IDT.

© 2015, 2016 Integrated DNA Technologies. All rights reserved. Trademarks contained herein are the property of Integrated DNA Technologies, Inc. or their respective owners. For specific trademark and licensing information, see www.idtdna.com/trademarks.


Predesigned qPCR Assays

Probe-based qPCR assays for quantification of human, mouse, and rat gene expression. Order in plates or tubes.

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