Identification of Valid Reference Genes for the Normalization of RT-qPCR Expression Studies in Human Breast Cancer Cell Lines Treated with and without Transient Transfection

CORC > 昆明动物研究所 > 昆明动物研究所 > 遗传资源与进化国家重点实验室 > 分子进化基因组学

	Identification of Valid Reference Genes for the Normalization of RT-qPCR Expression Studies in Human Breast Cancer Cell Lines Treated with and without Transient Transfection
	Liu LL 1,2; Zhao H 1; Ma TF 1; Ge F 2,3; Chen CS 3; Zhang YP[*]1,4
刊名	PLOS ONE
	2015
卷号	10 期号:1 页码:e0117058
通讯作者	zhangyp@mail.kiz.ac.cn
合作状况	其它
英文摘要	Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is a powerful technique for examining gene expression changes during tumorigenesis. Target gene expression is generally normalized by a stably expressed endogenous reference gene; however, reference gene expression may differ among tissues under various circumstances. Because no valid reference genes have been documented for human breast cancer cell lines containing different cancer subtypes treated with transient transfection, we identified appropriate and reliable reference genes from thirteen candidates in a panel of 10 normal and cancerous human breast cell lines under experimental conditions with/without transfection treatments with two transfection reagents. Reference gene expression stability was calculated using four algorithms (geNorm, NormFinder, BestKeeper and comparative delta Ct), and the recommended comprehensive ranking was provided using geometric means of the ranking values using the RefFinder tool. GeNorm analysis revealed that two reference genes should be sufficient for all cases in this study. A stability analysis suggests that 18S rRNA-ACTB is the best reference gene combination across all cell lines; ACTB-GAPDH is best for basal breast cancer cell lines; and HSPCB-ACTB is best for ER+ breast cancer cells. After transfection, the stability ranking of the reference gene fluctuated, especially with Lipofectamine 2000 transfection reagent in two subtypes of basal and ER+ breast cell lines. Comparisons of relative target gene (HER2) expression revealed different expressional patterns depending on the reference genes used for normalization. We suggest that identifying the most stable and suitable reference genes is critical for studying specific cell lines under certain circumstances.
收录类别	SCI
资助信息	This work was supported by grants of the National Natural Science Foundation of China (Grant Nos. 31060302 and 31260032) (http://www.nsfc.gov. cn), the Natural Science Foundation of Yunnan Province (Grant No. 2010CD010)
语种	英语
WOS记录号	WOS:000350336000037
公开日期	2015-03-27
内容类型	期刊论文
源URL	[http://159.226.149.42:8088/handle/152453/8287]
专题	昆明动物研究所_分子进化基因组学昆明动物研究所_动物模型与人类重大疾病机理重点实验室昆明动物研究所_遗传资源与进化国家重点实验室昆明动物研究所_肿瘤生物学
作者单位	1.Laboratory for Conservation and Utilization of Bio-resource, Yunnan University, Kunming, China 2.Department of Endocrine Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016, China 3.Key Laboratory of Animal Models and Human Disease Mechanisms of Chinese Academy of Sciences & Yunnan Province, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan, China 4.State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China
推荐引用方式 GB/T 7714	Liu LL,Zhao H,Ma TF,et al. Identification of Valid Reference Genes for the Normalization of RT-qPCR Expression Studies in Human Breast Cancer Cell Lines Treated with and without Transient Transfection[J]. PLOS ONE,2015,10(1):e0117058.
APA	Liu LL,Zhao H,Ma TF,Ge F,Chen CS,&Zhang YP[*].(2015).Identification of Valid Reference Genes for the Normalization of RT-qPCR Expression Studies in Human Breast Cancer Cell Lines Treated with and without Transient Transfection.PLOS ONE,10(1),e0117058.
MLA	Liu LL,et al."Identification of Valid Reference Genes for the Normalization of RT-qPCR Expression Studies in Human Breast Cancer Cell Lines Treated with and without Transient Transfection".PLOS ONE 10.1(2015):e0117058.