Profiling of microRNAs in mouse brain with real-time PCR array / 北京大学学报(医学版)

ABSTRACT

Objective:To examine global expression levels of microRNAs(miRNAs) in mouse cerebrum and to provide an important basis for detailed studies of individual miRNAs,their target genes,the miRNA-related regulatory networks in the mammalian central nervous system,and their implications in diseases.Methods:Low molecular weight RNA from cerebrum of five C57BL/6J mice were tailed and reverse transcribed by extended RT-primer.miRNA primers were carefully designed and arrayed on plates according to the Tm of each primer.PCR was carried out at different annealing temperatures using a gradient real-time PCR instrument.The relative expression level of each miRNA was calculated using 5sRNA for normalization.Results:Among the 285 miRNAs detected,260 were positive with varying abundance.Their frequency distribution was approximately a normal distribution.The expression levels of most miRNAs were in accordance with previously published results by microarray.However,the positive rate was higher than that detected by microarray.miRNAs originating from the same hairpin precursors expressed at similar or significantly different levels.Clusters of proximal miRNAs were similar or quite different in abundance.It is suggested that the fate of miRNA after transcription determined their abundance.Conclusion:Using the RNA-tailing and primer-extension PCR array method,we obtained expression profile of miRNA in mouse cerebrum,especially the relative expression data of many low abundant miRNA in mouse cerebrum,which will be of special help for studying the fine-tuning function of low-level miRNAs.