Target specific and non-biased gene amplification

TGene expression analysis is possible without changing copy number ratio in a sample!

Introduction

　In comprehensive gene expression analysis such as T cell receptor (TCR) repertoire analysis, it is important that target specific and non-biased gene amplification products are obtained. Conventional methods often ligate adapters with restriction enzyme site to synthesized cDNA ends and treat adapter-ligated cDNAs with said restriction enzyme before specific amplification. In such methods, however, unused and untreated adapters often inhibit target specific and non-biased amplification.　
　This invention performs target specific, non-biased gene amplification by combination of uracil-containing adapter and uracil DNA glycosylase (UNG) treatment (Fig 1).

Fig. 1. Illustration of this invention when used in TCR repertoire analysis

Fig. 2. Comparison of performance as viewed from TCR sequencing success

Effect

　TCR sequencing by NGS was more successfully performed with sufficient read length and overlaps among reads in TCR repertoire analysis using amplified products by this invention, when compared with that using products by conventional adaptor ligation method (Fig 2).
　This invention performs gene amplification without changes of copy number ratio in a sample, showing usefulness for various applications other than TCR analysis, such as RNA-seq, intestinal flora analysis and environmental (cell free) DNA analysis. The kit to do this invention is simply available by adding uracil-containing adapter and UNG to conventional PCR kit.

IP Data

IP No.　 ： WO2016/136716　(FR,GB,DE,JP,US)　　
Inventor ： OGASAWARA Koetsu
keyword ： TCR, Tcellreceptor, NGS