High-throughput SuperSAGE for digital gene expression analysis of multiple samples using next generation sequencing.

High-throughput SuperSAGE for digital gene expression analysis of multiple samples using next generation sequencing.
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DOI:
10.1371/journal.pone.0012010
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发表时间:
2010-08-06
期刊:
影响因子:
3.7
通讯作者:
Terauchi R
Terauchi R
中科院分区:
综合性期刊3区
文献类型:
--
作者:
Matsumura H;Yoshida K;Luo S;Kimura E;Fujibe T;Albertyn Z;Barrero RA;Krüger DH;Kahl G;Schroth GP;Terauchi R

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我们建立了SuperSAGE技术与下一代测序相结合的方案,创造了“高重复性(HT-)SuperSAGE”。SuperSAGE是一种数字基因表达谱分析方法,允许从表达的转录物中分离26-bp标签片段。在本方案中,采用索引(条形码)序列来区分来自不同样品的标签。这样的条形码允许研究人员通过简单地汇集文库,在单个测序运行中分析来自许多样品的转录组的数字标签。在这里,我们证明了HT-SuperSAGE提供了高度灵敏、可重复和准确的数字基因表达数据。通过增加在HT-SuperSAGE分析的吞吐量,可以预见到各种应用,并在本研究中提供了几个例子,包括分析激光显微切割的细胞,生物复制品和标签提取使用不同的锚定酶。
We established a protocol of the SuperSAGE technology combined with next-generation sequencing, coined “High-Throughput (HT-) SuperSAGE”. SuperSAGE is a method of digital gene expression profiling that allows isolation of 26-bp tag fragments from expressed transcripts. In the present protocol, index (barcode) sequences are employed to discriminate tags from different samples. Such barcodes allow researchers to analyze digital tags from transcriptomes of many samples in a single sequencing run by simply pooling the libraries. Here, we demonstrated that HT-SuperSAGE provided highly sensitive, reproducible and accurate digital gene expression data. By increasing throughput for analysis in HT-SuperSAGE, various applications are foreseen and several examples are provided in the present study, including analyses of laser-microdissected cells, biological replicates and tag extraction using different anchoring enzymes.
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