Selection and evaluation of stable housekeeping genes for gene expression normalization in carbon nanoparticle-induced acute pulmonary inflammation in mice

Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) is a highly specific and sensitive technique for the quantification of gene expression on the mRNA levels. But use of unconfirmed housekeeping genes (HKGs) could lead to misinterpretation of the expression of genes of interest (GOI). In this study, the stability and suitability of 11 frequently used housekeeping genes, namely 18S rRNA, ACTB, B2M, CYPA, GADPH, GUSB, HMBS, HPRT1, RPL13A, SDHA and TBP in 36 lung tissues isolated from either wild-type (WT) mice or p50 knock out (p50−/−) mice or p105 knock-out (p105−/−) mice which were treated with either carbon nanoparticle (CNP) or H2O or non-treated, have been validated by geNorm, NormFinder and BestKeeper programs. The expression levels of ACTB, GUSB and RPL13A were the most constant in lung tissues across three genotypes and three kinds of treatments. A set of three most stable genes is found sufficient to be used as housekeeping genes for lung tissues in studies of similar design.

Research highlights
► Stability of 11 frequently used housekeeping genes was analyzed.
► Expression levels of ACTB, GUSB and RPL13A were the most stable.
► Different expression level using different housekeeping genes.