Identification of suitable reference genes for gene expression measurement in uterine sarcoma and carcinosarcoma tumors

ObjectiveTo date no suitable reference genes have been identified in carcinosarcomas and non-epithelial malignant tumors of corpus uteri for normalizing real-time RT-PCR (qRT-PCR) assays. The purpose of this study was to select appropriate references for gene expression studies in these tumors.Materials and methodsWe used RNA extracts from 75 tissue samples, representing 50 tumors and 25 fragments of normal uterine tissues obtained from 50 patients treated for mixed tumors, smooth muscle sarcoma and stromal sarcoma of the uterus. qRT-PCR for five potential reference (housekeeping) genes, namely B2M, HMBS, HPRT1, TBP and UBC, was performed. The expression stability of these genes was assayed using geNorm software application.ResultsThe analysis of gene expression data with geNorm identified HPRT1 as the most stable reference gene, followed by UBC and HMBS, for all the investigated tissues. When stratified by disease, the results still pointed at HPRT1 as the gene that retained the greatest robustness in mixed tumors as well as in smooth muscle and stromal sarcomas.ConclusionsOur work is the first report on reference gene selection for qRT-PCR applications in mixed tumors, smooth muscle sarcoma and stromal sarcoma of the uterus. A ranking of candidate genes’ stability values for the three types of tumors is provided and might serve as a valuable guide for future gene expression studies of these rare entities.

► In this study we report on suitable reference genes for normalizing qRT-PCR assays. ► We analyzed mixed tumors, smooth muscle sarcomas and stromal sarcomas of the uterus. ► We examined five genes’ stability values for the three tumor types. ► HPRT1 was identified as the most stable reference gene, followed by UBC and HMBS. ► A ranking of reference genes is provided.