Deep mRNA Sequencing for In Vivo Functional Analysis of Cardiac Transcriptional Regulators Application to Gαq
Deep mRNA Sequencing for In Vivo Functional Analysis of Cardiac Transcriptional Regulators Application to Gαq
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DOI：
10.1161/circresaha.110.217513
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发表时间：
2010-05-14
影响因子：
20.1
通讯作者：
Dorn, Gerald W., II
中科院分区：
文献类型：
--
作者：
Matkovich, Scot J.;Zhang, Yan;Dorn, Gerald W., II
基本原理：转录谱可以检测亚临床心脏病，并提供疾病病因和功能状态的见解。目前的微阵列为基础的方法是昂贵的，并受到artifaction.Objective：开发RNA测序方法，使用下一代大规模并行平台的高通量全面分析个别小鼠心脏转录组。比较测序和阵列为基础的转录分析的结果，在良好的特点G α q转基因小鼠肥大/cardiomyopathy model.Methods和Results：的技术制备单独的条形码小鼠心脏RNA库的Illumina基因组分析仪II重测序。RNA测序显示，234个高丰度转录物（>60拷贝/细胞）占总心脏mRNA的55%。通过Illumina RNA测序和Affyssin Mouse Gene 1.0 ST阵列对G alpha q转基因和非转基因心脏进行的平行转录谱分析显示，mRNA表达的动态范围上级，并且通过RNA测序报告低丰度转录本的特异性增强。基因芯片和RNA测序高度丰富的转录本之间G α q和非转基因心脏的差异mRNA表达相关性良好。RNA测序是上级阵列准确定量低丰度的基因，这代表了大多数的G α q转基因model.Conclusions调控基因：RNA测序是快速，准确，灵敏的识别丰富和罕见的心脏转录，并具有显着的优势，在时间和成本效益的微阵列分析。(Circ Res. 2010; 106：1459-1467）。
Rationale: Transcriptional profiling can detect subclinical heart disease and provide insight into disease etiology and functional status. Current microarray-based methods are expensive and subject to artifact.Objective: To develop RNA sequencing methodologies using next generation massively parallel platforms for high throughput comprehensive analysis of individual mouse cardiac transcriptomes. To compare the results of sequencing- and array-based transcriptional profiling in the well-characterized G alpha q transgenic mouse hypertrophy/cardiomyopathy model.Methods and Results: The techniques for preparation of individually bar-coded mouse heart RNA libraries for Illumina Genome Analyzer II resequencing are described. RNA sequencing showed that 234 high-abundance transcripts (>60 copies/cell) comprised 55% of total cardiac mRNA. Parallel transcriptional profiling of G alpha q transgenic and nontransgenic hearts by Illumina RNA sequencing and Affymetrix Mouse Gene 1.0 ST arrays revealed superior dynamic range for mRNA expression and enhanced specificity for reporting low-abundance transcripts by RNA sequencing. Differential mRNA expression in G alpha q and nontransgenic hearts correlated well between microarrays and RNA sequencing for highly abundant transcripts. RNA sequencing was superior to arrays for accurately quantifying lower-abundance genes, which represented the majority of the regulated genes in the G alpha q transgenic model.Conclusions: RNA sequencing is rapid, accurate, and sensitive for identifying both abundant and rare cardiac transcripts, and has significant advantages in time-and cost-efficiencies over microarray analysis. (Circ Res. 2010; 106:1459-1467.)