Next generation sequencing (NGS) has been the driving force in molecular biology and biomedicine. After completion of the human genome project, this technique arose and is now used to tackle the questions remaining after identification of the human genetic information. We have to answer the terms of regulation and transcriptional importance of specific genes. The method, where sequencing and gene expression come together is RNA-Seq. This technique has as all methods benefits and shortcomings, but in combination with quantitative PCR (qPCR) and gene specific sequencing it is a powerful tool that can and will alter our procedure of analyzing patient samples. Possible pitfalls and solutions are provided to tackle problems not only for RNA-Seq but also for CNV (copy number variation) analysis, which mostly employ qPCR. Since oncology is a major field of interest and represents a good example as research area due to a multitude of starting material, it has been chosen to be the focus of this methodological overview.
- Loewe RP. (2012) Combinational Usage of Next Generation Sequencing and qPCR for the analysis of tumour samples. Methods [Epub ahead of print]. [abstract]