Reverse Transcription Free Microarray Analysis

描述（由申请人提供）：该项目的目标是开发一种新的微阵列系统，用于直接用总RNA进行基因表达分析，而无需标记和反转录到cDNA。实现这一目标将为微阵列领域提供一个关键的突破，并建立新的标准。事实上，尽管微阵列技术已经被广泛研究并在许多应用中被证明是成功的，但在这一领域仍然存在许多挑战。首先，传统的微阵列检测需要逆转录总RNA并用报告分子标记，成本高，耗时长。其次，目标序列的PCR扩增可能产生系统误差和杂交信号偏差。第三，微阵列仪器的成本很高。降低仪器和日常操作的成本将使小型研究机构和诊所更容易使用微阵列技术。本项目的关键创新点是利用靶分子与纳米级粒子的静电相互作用来检测RNA的新技术。该检测技术允许直接使用总RNA进行杂交实验，不需要掺入报告基因，也不需要从RNA逆转录到cDNA。该技术还允许廉价的仪器用于基因表达分析中DNA和RNA的高灵敏度检测。最近有报道称，在肿瘤发生和化疗预后的临床试验中使用微阵列（Science, 303:1754），仅在美国就有500多家临床实验室和数百家生命科学研究实验室迅速采用了一种先进且廉价的微阵列平台。

DESCRIPTION (provided by applicant): The goal of this project is to develop a new microarray system for performing gene expression analysis directly with total RNA without labeling and reverse transcription to cDNA. Achieving this goal will provide a critical breakthrough and establish a new standard in microarray field. Indeed, although microarray technique has been widely studied and proved to be successful in many applications, there are still many challenges in this field. First, the traditional microarray detection requires the total RNA to be reverse transcribed and labeled with reporter molecules, which is costly and time-consuming. Second, the PCR amplification of target sequences may give rise to systematic errors and biased hybridization signals. Third, the cost for a microarray instrumentation is high. Reducing the cost of instrumentation and day-by-day operation will make microarray technology more accessible to small research institutions and clinics. The key innovation of this project is a new RNA detection technique which employs electrostatic interaction of the target molecules and nanometer-size particles. This detection technique allows to use total RNA directly for hybridization experiment without reporter incorporation and reverse transcription from RNA to cDNA. The technique also allows inexpensive instrumentation to be used for highly-sensitive detection of DNA and RNA in gene expression analysis. With the recent reports of using microarrays in clinical tests for tumorogenesis and prognosis of a chemotherapy outcome (Science, 303:1754), introducing an advanced and inexpensive microarray platform will find fast adoption in more than 500 clinical laboratories and hundreds of life science research laboratories in US alone.