Highly sensitive and specific microRNA expression profiling using BeadArray technology

被引:75
作者
Chen, Jing [1 ]
Lozach, Jean [1 ]
Wickham, Eliza [1 ]
Barnes, Garcia Bret [1 ]
Luo, Shujun [1 ]
Mikoulitch, Ivan [1 ]
Zhou, Lixin [1 ]
Schroth, Gary [1 ]
Fan, Jian-Bing [1 ]
机构
[1] Illumina Inc, San Diego, CA 92121 USA
关键词
D O I
10.1093/nar/gkn387
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We have developed a highly sensitive, specific and reproducible method for microRNA (miRNA) expression profiling, using the BeadArray technology. This method incorporates an enzyme-assisted specificity step, a solid-phase primer extension to distinguish between members of miRNA families. In addition, a universal PCR is used to amplify all targets prior to array hybridization. Currently, assay probes are designed to simultaneously analyse 735 well-annotated human miRNAs. Using this method, highly reproducible miRNA expression profiles were generated with 100-200 ng total RNA input. Furthermore, very similar expression profiles were obtained with total RNA and enriched small RNA species (R-2 >= 0.97). The method has a 3.5-4 log (10(5)-10(9) molecules) dynamic range and is able to detect 1.2- to 1.3-fold-differences between samples. Expression profiles generated by this method are highly comparable to those obtained with RTPCR (R-2 = 0.850.90) and direct sequencing (R = 0.870.89). This method, in conjunction with the 96-sample array matrix should prove useful for high-throughput expression profiling of miRNAs in large numbers of tissue samples.
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页数:9
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