筛选LPS诱导下THP-1细胞稳定内参基因的研究
首发时间:2012-12-19
摘要:目的 筛选LPS诱导THP-1细胞前后稳定的内参基因。 方法 运用RT-qPCR方法,分析10个候选内参基因(ACTB、GAPDH、RPL37A、PPIB、PGK1、PPIA、SDHA、TBP、HPRT1、 RPL13A )在LPS刺激THP-1细胞前后不同时间的表达稳定性。 结果 经NormFinder和GeNorm软件分析可知,GAPDH、PGK1表达稳定,可用来准确校正定量结果;后期实验中同时选用GAPDH、PGK1有助于得到更可靠的结果。结论 RT-qPCR实验应该根据细胞和组织的类型及不同实验条件选择最合适的内参基因;选择2个或2个以上的内参基因将有助于得到更可靠的结果。
关键词: RT-PCR 内参基因 LPS THP-1细胞 NormFinder软件 GeNorm软件
For information in English, please click here
Gene expression results in lipopolysaccharide-stimulated THP-1 cells depend significantly on the choice of reference genes
Abstract:The choice of internal control genes is important since it may affect the study outcome in RT-qPCR. Indeed, it is well-known that expression levels of traditional reference genes can vary across tissue types and across experimental settings within one specific tissue type. The aim of this study is an evaluation of a set of housekeeping genes (HKGs) to be used in the normalization of gene expression in vitro different cultured cells, THP-1. The transcriptional stability of ten potential reference genes (ACTB,GAPDH,RPL37A,PPIB,PGK1,PPIA,SDHATBP, HPRT1 and RPL13A) were evaluated using RT-qPCR and were compared in different treatment, that was un-stimulated or LPS-stimulated cells. The raw Ct values were determined for each candidate gene at different time points following LPS-stimulated or unstimulated cells. Furthermore, all data were analyzed by the geNorm and NormFinder validation programs. Results indicated that DAPDH and PGK1 were the most stable internal control genes in this study. This study provides the comprehensive reported assessment of internal control genes for use in expression studies in vitro cultured cells. These findings further emphasize the need to accurately validate candidate internal control genes in the study before use in gene expression studies using RT-qPCR.
Keywords: RT-qPCR Reference genes LPS THP-1 NormFinder GeNorm
基金:
论文图表:
引用
No.****
同行评议
共计0人参与
勘误表
筛选LPS诱导下THP-1细胞稳定内参基因的研究
评论
全部评论0/1000