目的:寻找唐氏综合征模型鼠Ts65Dn和对照二倍体组小鼠海马组织中差异表达的microRNAs。方法:从小鼠的海马组织中提取小分子RNA,采用RNA加尾和引物延伸RT-PCR法检测microRNAs。通过geNorm软件和Normfinder软件分析选择最佳的内参照基因,将每种microRNAs按其引物Tm值排成阵列,用梯度实时PCR仪在不同的退火温度扩增,并计算每种相对于microRNAs内参照基因的表达量。结果:geNorm和Normfinder两种方法均选定miR-27a为最稳定的内参照基因,共检测了52种microRNAs,其中miR-33和miR-19a在Ts65Dn小鼠海马组织中表达显著下调,miR-130在Ts65Dn小鼠海马组织中表达显著上调,小鼠16号染色体三体区段包含的miR-802在Ts65Dn小鼠和对照二倍体小鼠海马组织中表达量均低,差异没有统计学意义。结论:MicroRNAs在Ts65Dn小鼠海马组织中的异常表达可能与唐氏综合征脑发育不良有关。
Objective:To identify the differentially expressed microRNAs in the hippocampus of Down Syndrome model mouse Ts65Dn.Methods: Low molecular weight RNA from hippocampus were tailed and reverse transcribed by extended RT-primer.MicroRNAs primers were arrayed on plates according to the Tm of each primer.PCR were carried out at different annealing temperatures using a gradient real-time PCR instrument.The relative expression level of each microRNAs was calculated using the most stable reference gene as normalizer which was selected by geNorm and Normfinder software.Results: miR-27a was identified as the most stable reference gene by geNorm and Normfinder software.Among the 52 microRNAs detected by real time PCR array,miR-33 and miR-19a were significantly down-regulated,whereas miR-130 were significantly up-regulated in the hippocampus of Ts65Dn mice as compared with the euploid control mice.The expression of miR-802,which was the trisomy chromosome 16 derived microRNAs,was very low in hippocampus with no difference between the two groups.Conclusion: MicroRNAs are dysregulated in the hippocampus of Ts65Dn mice,which may contribute to the reduced neurogenesis of Down syndrome.