Work

Application of Reverse Transcription-PCR and Real-Time PCR in Nanotoxicity Research

Year: 2012

Type: book-chapter

Abstract: Reverse transcription-polymerase chain reaction (RT-PCR) is a relatively simple and inexpensive technique to determine the expression level of target genes and is widely used in biomedical science research including nanotoxicology studies for semiquantitative analysis. Real-time PCR allows for the detection of PCR amplification in the exponential growth phase of the reaction and is much more quantitative than traditional RT-PCR. Although a number of kits and reagents for RT-PCR and real-time PCR are commercially available, the basic principles are the same. Here, we describe the procedures for total RNA isolation by using TRI Reagent, for reverse transcription (RT) by M-MLV reverse transcriptase, and for PCR by GoTaq(®) DNA Polymerase. And real-time PCR will be performed on an iQ5 multicolor real-time PCR detection system by using iQ™ SYBR Green Supermix.

Source: Methods in molecular biology

DOI: 10.1007/978-1-62703-002-1_7

Authors: Yiqun Mo, Rong Wan, Qunwei Zhang

Institution: University of Louisville

Language: English

Topic: Molecular Biology Techniques and Applications

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: green

Funders: American Lung Association, University of Louisville, National Institute of Environmental Health Sciences

Awards: G3297420083, UOFL ENVIRONMENTAL HEALTH SCIENCES TRAINING PROGRAM