Establishment of a reporter system to monitor silencing status in induced pluripotent stem cell lines

被引：19

作者：

Donai, Kenichiro ^{[1
]}

Kuroda, Kengo ^{[1
]}

Guo, Yijie ^{[1
]}

So, Kyoung-Ha ^{[1
]}

Sone, Hideko ^{[2
]}

Kobayashi, Masayuki ^{[3
]}

Nishimori, Katsuhiko ^{[1
]}

Fukuda, Tomokazu ^{[1
]}

机构：

[1] Tohoku Univ, Grad Sch Agr Sci, Aoba Ku, Sendai, Miyagi 9818555, Japan

[2] Natl Inst Environm Studies, Ctr Environm Risk Res, Hlth Risk Res Sect, Tsukuba, Ibaraki 3058506, Japan

[3] Akita Prefectural Univ, Grad Sch Bioresource Sci, Akita 0100195, Japan

来源：

ANALYTICAL BIOCHEMISTRY | 2013年 / 443卷 / 01期

基金：

日本学术振兴会;

关键词：

Induced pluripotent stem cells; Gene silencing; Reprogramming; Reporter system; TRANSCRIPTIONAL NETWORK; REPROGRAMMING FACTORS; LENTIVIRAL VECTORS; EXPRESSION; FIBROBLASTS; GENE; DIFFERENTIATION; GENERATION; INDUCTION; CLONING;

D O I：

10.1016/j.ab.2013.08.014

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Induced pluripotent stem (iPS) cells have proven to be an effective technology in regenerative medicine; however, the low efficiency of reprogramming is a major obstacle to the successful generation of iPS cell lines. One of the most important characteristics of a high-quality iPS cell line is the inactivation of transgenes driven by a retrovirus-derived long terminal repeat promoter. In this study, we established a novel marker system containing three kinds of proteins: secreted-type luciferase (MetLuc), copepod Pontellina plumate green fluorescent protein (copGFP), and an antibiotic-resistant gene product (Neo(r)). The introduction of MetLuc-copGFP-Neo(r) in mouse embryonic fibroblasts (MEFs) allowed us to monitor the reporter expression changes as an indicator of the state of silencing during reprogramming. Transformation of iPS cells induced a remarkable reduction in reporter activity, indicating that the retroviral silencing was detected successfully. Our system enables us to monitor the silencing status of transgenes and to efficiently select iPS cell lines that can be used for further applications. (C) 2013 Elsevier Inc. All rights reserved.

引用

页码：104 / 112

页数：9

共 27 条

[1] Conversion of Ancestral Fibroblasts to Induced Pluripotent Stem Cells [J].