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微电泳阵列设备上核酸检测的规模化:从少于十个转录本进行高动态范围多基因读出。

Scaling of nucleic acid assays on microelectrophoresis array devices: high-dynamic range multi-gene readout from less than ten transcripts.

作者信息

Ueberfeld Joern, Ehrlich Daniel J

机构信息

Whitehead Institute for Biomedical Research Nine Cambridge Center, Cambridge, MA, USA.

出版信息

Electrophoresis. 2009 Jun;30(12):2090-9. doi: 10.1002/elps.200800774.

Abstract

In this paper we describe progress in using the prodigious data-collecting ability of multilane microelectrophoresis instruments to bear on problems in scaled nucleic acid assays. We emphasize compound stacking and solid-support loading as means to concentrate <100 pg samples for direct injection. Reaction Mapping is applied to readout quantitative polymerase chain reaction gene-expression and as a way to practically overcome difficulty in interpreting amplification curves of multiplexed quantitative polymerase chain reaction at 20-50 gene/well complexity. We demonstrate multiplexed readout of gene expression over an abundancy range of 9 Log 2 units starting with reverse-transcribed samples as small as five molecules in each sample.

摘要

在本文中,我们描述了利用多通道微电泳仪器强大的数据收集能力来解决核酸分析规模化问题所取得的进展。我们着重介绍了复合堆积和固相载体加载,以此作为浓缩小于100 pg样品以进行直接进样的方法。反应图谱被应用于读取定量聚合酶链反应基因表达,并作为一种切实克服在20 - 50个基因/孔复杂度下解释多重定量聚合酶链反应扩增曲线困难的方法。我们展示了从每个样品仅五个分子的逆转录样品开始,在9个Log₂单位的丰度范围内对基因表达进行多重读出。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e2c6/2760978/1403cf51a28e/nihms147354f1.jpg

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