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Sleeping beauty transposon-mediated poly(A)-trapping and insertion mutagenesis in mouse embryonic stem cells
Zhao, Yi1; Song, Guili2; Ren, Jing2; Li, Qing2; Zhong, Shan1,3,4; Cui, Zongbin2
2018-10-01
Source PublicationENVIRONMENTAL AND MOLECULAR MUTAGENESIS
ISSN0893-6692
Volume59Issue:8Pages:687-697
Abstract

Saturation mutagenesis of all endogenous genes within the mouse genome remains a challenging task, although a plenty of gene-editing approaches are available for this purpose. Here, a poly(A)-trap vector was generated for insertion mutagenesis in mouse embryonic stem (mES) cells. This vector contains an expression cassette of neomycin (Neo)-resistant gene lacking a poly(A) signal and flanked by two inverted terminal repeats of the Sleeping Beauty (SB) transposon. The whole poly(A)-trap cassette can transpose into target TA dinucleotides, properly splice with endogenous genes and effectively interrupt the transcription of trapped genes in mES cells after transient induction of SB expression by doxycycline (DOX)-treatment at 1 g/ml, leading to the formation of multiple geneticin (G418)-resistant cell clones. In the first round of mutation screening, we identified six transposition events from 23 cell clones, including four inserted into an endogenous gene and two landed between endogenous genes. The abilities of self-renewal, totipotency, genetic stability and differentiation of syngap1(+/-) cells were not affected by DOX-treatment and G418-selection. These findings suggest that this SB transposon-mediated poly(A)-trap vector can be used as an alternative tool for a large-scale screening of mES cells with a gene mutation and for further generation of mutant mouse strains. Environ. Mol. Mutagen. 59:687-697, 2018. (c) 2018 Wiley Periodicals, Inc.

Keywordmouse embryonic stem cells sleeping beauty transposon poly(A) trapping insertion mutagenesis
DOI10.1002/em.22234
Indexed BySCI
Language英语
WOS Research AreaEnvironmental Sciences & Ecology ; Genetics & Heredity ; Toxicology
WOS SubjectEnvironmental Sciences ; Genetics & Heredity ; Toxicology
WOS IDWOS:000447275100002
WOS KeywordZINC-FINGER NUCLEASES ; GENE-FUNCTION ; HUMAN GENOME ; EXPRESSION ; SYSTEM ; ESTABLISHMENT ; TRANSCRIPTION ; ENDONUCLEASE ; INTEGRATION ; EFFECTORS
PublisherWILEY
Citation statistics
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/29264
Collection水生生物分子与细胞生物学研究中心_期刊论文
Corresponding AuthorZhong, Shan; Cui, Zongbin
Affiliation1.Wuhan Univ, Sch Basic Med Sci, Dept Genet, Wuhan, Hubei, Peoples R China
2.Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan, Hubei, Peoples R China
3.Hubei Prov Key Lab Dev Originated Dis, Wuhan, Hubei, Peoples R China
4.Hubei Prov Key Lab Allergy & Immunol, Wuhan, Hubei, Peoples R China
Recommended Citation
GB/T 7714
Zhao, Yi,Song, Guili,Ren, Jing,et al. Sleeping beauty transposon-mediated poly(A)-trapping and insertion mutagenesis in mouse embryonic stem cells[J]. ENVIRONMENTAL AND MOLECULAR MUTAGENESIS,2018,59(8):687-697.
APA Zhao, Yi,Song, Guili,Ren, Jing,Li, Qing,Zhong, Shan,&Cui, Zongbin.(2018).Sleeping beauty transposon-mediated poly(A)-trapping and insertion mutagenesis in mouse embryonic stem cells.ENVIRONMENTAL AND MOLECULAR MUTAGENESIS,59(8),687-697.
MLA Zhao, Yi,et al."Sleeping beauty transposon-mediated poly(A)-trapping and insertion mutagenesis in mouse embryonic stem cells".ENVIRONMENTAL AND MOLECULAR MUTAGENESIS 59.8(2018):687-697.
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