High-Throughput SuperSAGE for Digital Gene Expression Analysis of Multiple Samples Using Next Generation Sequencing

PLoS One

Detlev H. Krüger¹  Roberto A. Barrero¹  Takahiro Fujibe²  Kentaro Yoshida³  Eiji Kimura⁴  Ryohei Terauchi⁴  Günter Kahl⁵  Hideo Matsumura⁶  Shujun Luo⁶  Zayed Albertyn⁶  Gary P. Schroth⁷ 

[1] Centre for Comparative Genomics, Murdoch University, Perth, Australia;Department of Anatomy, Iwate Medical University, Morioka, Japan;Gene Research Center, Shinshu University, Ueda, Japan;Illumina, Inc., Hayward, California, United States of America;Institute of Medical Virology, University Hospital Charite, Berlin, Germany;Iwate Biotechnology Research Center, Kitakami, Japan;Molecular BioSciences, Biocentre, University of Frankfurt am Main, Frankfurt, Germany

关键词: Rice;    Gene expression;    Arabidopsis thaliana;    Polymerase chain reaction;    Sequence tagged site analysis;    Expressed sequence tags;    Sequence assembly tools;    Sequence databases;   

DOI  :  10.1371/journal.pone.0012010

学科分类：医学（综合）

来源: Public Library of Science

【 摘 要 】

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.

【 授权许可】

CC BY