Targeted RNA sequencing enhances gene expression profiling of ultra-low input samples.
Targeted RNA sequencing enhances gene expression profiling of ultra-low input samples.
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靶向RNA测序增强了超低输入样品的基因表达分析。
DOI：
10.1080/15476286.2020.1777768
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发表时间：
2020-12
期刊：
影响因子：
4.1
通讯作者：
Clark MB
中科院分区：
文献类型：
--
作者：
Curion F;Handel AE;Attar M;Gallone G;Bowden R;Cader MZ;Clark MB
RNA-seq 是许多生物系统中基因表达分析的标准方法。由于转录组的宽动态范围和复杂性，RNA-seq 提供了不完整的表征，尤其是低表达基因和转录本。靶向 RNA 测序 (RNA CaptureSeq) 重点对感兴趣的基因进行测序，为转录本检测和定量提供极高的灵敏度。然而，CaptureSeq 的使用主要集中在大量样本上，其在极小细胞群中的性能尚不清楚。在这里，我们展示了 CaptureSeq 极大地增强了超低输入样品中目标基因的转录组分析，并提供了与批量样品相同的性能。我们使用多个探针组对 iPSC 衍生的皮质神经元样本验证 CaptureSeq 的性能。我们展示了目标基因高达 275 倍的富集、10% 额外基因的检测以及已识别基因亚型的增加超过 5 倍。对掺入对照的分析表明，CaptureSeq 提高了检测灵敏度和表达定量。与大脑皮层发育 CORTECON 数据库的比较显示，CaptureSeq 增强了样本分化阶段的识别。 CaptureSeq 对来自超低输入样本的数百至数千个目标基因提供灵敏、可靠和定量的表达测量，并且有可能在样本有限时极大地增强转录组分析。
RNA-seq is the standard method for profiling gene expression in many biological systems. Due to the wide dynamic range and complex nature of the transcriptome, RNA-seq provides an incomplete characterization, especially of lowly expressed genes and transcripts. Targeted RNA sequencing (RNA CaptureSeq) focuses sequencing on genes of interest, providing exquisite sensitivity for transcript detection and quantification. However, uses of CaptureSeq have focused on bulk samples and its performance on very small populations of cells is unknown. Here we show CaptureSeq greatly enhances transcriptomic profiling of target genes in ultra-low-input samples and provides equivalent performance to that on bulk samples. We validate the performance of CaptureSeq using multiple probe sets on samples of iPSC-derived cortical neurons. We demonstrate up to 275-fold enrichment for target genes, the detection of 10% additional genes and a greater than 5-fold increase in identified gene isoforms. Analysis of spike-in controls demonstrated CaptureSeq improved both detection sensitivity and expression quantification. Comparison to the CORTECON database of cerebral cortex development revealed CaptureSeq enhanced the identification of sample differentiation stage. CaptureSeq provides sensitive, reliable and quantitative expression measurements on hundreds-to-thousands of target genes from ultra-low-input samples and has the potential to greatly enhance transcriptomic profiling when samples are limiting.
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影响因子：
14.9
作者：
Frankish A;Diekhans M;Ferreira AM;Johnson R;Jungreis I;Loveland J;Mudge JM;Sisu C;Wright J;Armstrong J;Barnes I;Berry A;Bignell A;Carbonell Sala S;Chrast J;Cunningham F;Di Domenico T;Donaldson S;Fiddes IT;García Girón C;Gonzalez JM;Grego T;Hardy M;Hourlier T;Hunt T;Izuogu OG;Lagarde J;Martin FJ;Martínez L;Mohanan S;Muir P;Navarro FCP;Parker A;Pei B;Pozo F;Ruffier M;Schmitt BM;Stapleton E;Suner MM;Sycheva I;Uszczynska-Ratajczak B;Xu J;Yates A;Zerbino D;Zhang Y;Aken B;Choudhary JS;Gerstein M;Guigó R;Hubbard TJP;Kellis M;Paten B;Reymond A;Tress ML;Flicek P
通讯作者：
Flicek P
影响因子：
7
作者：
Cieslik M;Chugh R;Wu YM;Wu M;Brennan C;Lonigro R;Su F;Wang R;Siddiqui J;Mehra R;Cao X;Lucas D;Chinnaiyan AM;Robinson D
通讯作者：
Robinson D
影响因子：
14.9
作者：
Halvardson J;Zaghlool A;Feuk L
通讯作者：
Feuk L
影响因子：
6.4
作者：
Amorim-Vaz, Sara;Tran, Van Du T.;Sanglard, Dominique
通讯作者：
Sanglard, Dominique
影响因子：
6.8
作者：
AKAIKE, H
通讯作者：
AKAIKE, H