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Evaluation of a new high-dimensional miRNA profiling platform

机译：评估新的高维miRNA分析平台

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Background MicroRNAs (miRNAs) are a class of approximately 22 nucleotide long, widely expressed RNA molecules that play important regulatory roles in eukaryotes. To investigate miRNA function, it is essential that methods to quantify their expression levels be available. Methods We evaluated a new miRNA profiling platform that utilizes Illumina's existing robust DASL chemistry as the basis for the assay. Using total RNA from five colon cancer patients and four cell lines, we evaluated the reproducibility of miRNA expression levels across replicates and with varying amounts of input RNA. The beta test version was comprised of 735 miRNA targets of Illumina's miRNA profiling application. Results Reproducibility between sample replicates within a plate was good (Spearman's correlation 0.91 to 0.98) as was the plate-to-plate reproducibility replicates run on different days (Spearman's correlation 0.84 to 0.98). To determine whether quality data could be obtained from a broad range of input RNA, data obtained from amounts ranging from 25 ng to 800 ng were compared to those obtained at 200 ng. No effect across the range of RNA input was observed. Conclusion These results indicate that very small amounts of starting material are sufficient to allow sensitive miRNA profiling using the Illumina miRNA high-dimensional platform. Nonlinear biases were observed between replicates, indicating the need for abundance-dependent normalization. Overall, the performance characteristics of the Illumina miRNA profiling system were excellent.
机译：背景技术MicroRNA（miRNA）是一类大约22个核苷酸长的，广泛表达的RNA分子，在真核生物中起重要的调节作用。为了研究miRNA的功能，必须有量化其表达水平的方法。方法我们评估了一个新的miRNA分析平台，该平台利用了Illumina现有的强大DASL化学作为测定的基础。使用来自五名结肠癌患者和四个细胞系的总RNA，我们评估了重复和不同输入RNA量下miRNA表达水平的可重复性。 Beta测试版本由Illumina的miRNA分析应用程序中的735个miRNA靶组成。结果板内样品重复试验之间的重复性良好（Spearman相关性0.91至0.98），板对板重复性重复试验在不同天进行（Spearman相关性0.84至0.98）。为了确定是否可以从广泛的输入RNA中获得质量数据，将25 ng至800 ng范围内的数据与200 ng范围内的数据进行了比较。在整个RNA输入范围内均未观察到影响。结论这些结果表明，使用Illumina miRNA高维平台，很少量的起始材料即可进行敏感的miRNA分析。在复制之间观察到非线性偏差，表明需要依赖于丰度的归一化。总体而言，Illumina miRNA分析系统的性能特征非常出色。

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