オカノ　テルオ   OKANO Teruo   岡野　光夫    所属   研究施設 研究施設    職種   特任顧問
論文種別	原著
言語種別	英語
査読の有無	査読あり
表題	β2-Microglobulin is an appropriate reference gene for RT-PCR-based gene expression analysis of hematopoietic stem cells.
掲載誌名	正式名：Regenerative therapy 略　 称：Regen Ther ISSNコード：23523204/23523204
掲載区分	国外
巻・号・頁	1,pp.91-97
著者・共著者	MATSUZAKI Yu†, UMEMOTO Terumasa*, TANAKA Yuji, OKANO Teruo, YAMATO Masayuki*
発行年月	2015/06
概要	Real-time reverse transcription polymerase chain reaction (RT-PCR) is regarded as one of the most useful and powerful tools for characterizing hematopoietic stem cells (HSCs), because samples of extremely small cell numbers can be analyzed. The expression levels determined by RT-PCR are based on relative quantification; therefore, the selection of an appropriate reference gene with a relatively stable expression level under most conditions is crucial. Here, we determined that beta2-microglobulin (B2m) is an appropriate reference gene for analyzing mouse HSCs by a novel method using single-cell RT-PCR. Clonally sorted HSCs were subjected to RT reactions with exogenous RNA fragments and then to real-time PCR. Next, the relative gene expression levels of 4 well-known housekeeping genes were quantified in each single cell sample based on the threshold cycle of exogenous RNA. The analysis revealed that B2m expression was reproducibly detected in almost all HSCs and that B2m had the most stable expression level among the compared genes, even after the cells had been cultured under various conditions. Thus, our results indicate that B2m can reliably be used as a reference gene for the relative quantification of expression levels in HSCs across various conditions. Furthermore, our work proposes a novel method for the selection of appropriate reference genes.
DOI	10.1016/j.reth.2015.04.003
PMID	31245448