Global RT-PCR and RT-qPCR Analysis of the mRNA Expression of the Human PTPome

Methods Mol Biol. 2016:1447:25-37. doi: 10.1007/978-1-4939-3746-2_2.

Abstract

Comprehensive comparative gene expression analysis of the tyrosine phosphatase superfamily members (PTPome) under cell- or tissue-specific growth conditions may help to define their individual and specific role in physiology and disease. Semi-quantitative and quantitative PCR are commonly used methods to analyze and measure gene expression. Here, we describe technical aspects of PTPome mRNA expression analysis by semi-quantitative RT-PCR and quantitative RT-PCR (RT-qPCR). We provide a protocol for each method consisting in reverse transcription followed by PCR using a global platform of specific PTP primers. The chapter includes aspects from primer validation to the setup of the PTPome RT-qPCR platform. Examples are given of PTP-profiling gene expression analysis using a human breast cancer cell line upon long-term or short-term treatment with cell signaling-activation agents.

Keywords: PTPome; Protein tyrosine phosphatase; Real-time quantitative PCR; Reverse transcription PCR.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Gene Expression Profiling / methods*
  • Humans
  • MCF-7 Cells
  • Protein Tyrosine Phosphatases / genetics*
  • RNA, Messenger / genetics*
  • Real-Time Polymerase Chain Reaction / methods*
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • Transcriptome

Substances

  • RNA, Messenger
  • Protein Tyrosine Phosphatases