Home LiteratureArticle Details
PMID: 11258481 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Genome-wide insertional mutagenesis in human cells by the Drosophila mobile element Minos.

EMBO reports ·Vol. 1 ·No. 5 ·2000-11-00 ·Pages 416-21

Klinakis AG, Zagoraiou L, Vassilatis DK, Savakis C

Abstract

The development of efficient non-viral methodologies for genome-wide insertional mutagenesis and gene tagging in mammalian cells is highly desirable for functional genomic analysis. Here we describe transposon mediated mutagenesis (TRAMM), using naked DNA vectors based on the Drosophila hydei transposable element Minos. By simple transfections of plasmid Minos vectors in HeLa cells, we have achieved high frequency generation of cell lines, each containing one or more stable chromosomal integrations. The Minos-derived vectors insert in different locations in the mammalian genome. Genome-wide mutagenesis in HeLa cells was demonstrated by using a Minos transposon containing a lacZ-neo gene-trap fusion to generate a HeLa cell library of at least 10(5) transposon insertions in active genes. Multiple gene traps for six out of 12 active genes were detected in this library. Possible applications of Minos-based TRAMM in functional genomics are discussed.

MeSH Terms
Animals Base Sequence Cloning, Molecular DNA Transposable Elements DNA, Complementary/metabolism Drosophila melanogaster/genetics Exons Gene Library Genetic Techniques Genetic Vectors Genome, Human HeLa Cells Humans Models, Genetic Molecular Sequence Data Mutagenesis, Insertional Plasmids/metabolism Reverse Transcriptase Polymerase Chain Reaction Transfection
Chemicals
DNA Transposable Elements DNA, Complementary
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Klinakis A G
Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology, Hellas, Heraklion, Greece.
Zagoraiou L
Vassilatis D K
Savakis C
References (31)
31 references, click to expand
  1. Stable germline transformation of the malaria mosquito Anopheles stephensi.
    Nature. 2000 Jun 22;405(6789):959-62 PMID: 10879538
  2. Somatic integration and long-term transgene expression in normal and haemophilic mice using a DNA transposon system.
    Nat Genet. 2000 May;25(1):35-41 PMID: 10802653
  3. Extrachromosomal transposition of the transposable element Minos occurs in embryos of the silkworm Bombyx mori.
    Insect Mol Biol. 2000 Jun;9(3):277-81 PMID: 10886411
  4. Mouse embryonic stem cells and reporter constructs to detect developmentally regulated genes.
    Science. 1989 Apr 28;244(4903):463-5 PMID: 2497519
  5. Searching for pattern and mutation in the Drosophila genome with a P-lacZ vector.
    Genes Dev. 1989 Sep;3(9):1273-87 PMID: 2558049
  6. P-element-mediated enhancer detection: a versatile method to study development in Drosophila.
    Genes Dev. 1989 Sep;3(9):1288-300 PMID: 2558050
  7. P-element-mediated enhancer detection: an efficient method for isolating and characterizing developmentally regulated genes in Drosophila.
    Genes Dev. 1989 Sep;3(9):1301-13 PMID: 2558051
  8. Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice.
    Genes Dev. 1991 Sep;5(9):1513-23 PMID: 1653172
  9. Minos, a new transposable element from Drosophila hydei, is a member of the Tc1-like family of transposons.
    Nucleic Acids Res. 1991 Dec 11;19(23):6646 PMID: 1661410
  10. Mobile Minos elements from Drosophila hydei encode a two-exon transposase with similarity to the paired DNA-binding domain.
    Proc Natl Acad Sci U S A. 1994 May 24;91(11):4746-50 PMID: 8197129
  11. A large-scale gene-trap screen for insertional mutations in developmentally regulated genes in mice.
    Genetics. 1995 Feb;139(2):889-99 PMID: 7713439
  12. Capturing genes encoding membrane and secreted proteins important for mouse development.
    Proc Natl Acad Sci U S A. 1995 Jul 3;92(14):6592-6 PMID: 7604039
  13. Introduction of the transposable element Minos into the germ line of Drosophila melanogaster.
    Proc Natl Acad Sci U S A. 1995 Oct 10;92(21):9485-9 PMID: 7568159
  14. cRACE: a simple method for identification of the 5' end of mRNAs.
    Nucleic Acids Res. 1995 Sep 25;23(18):3796-7 PMID: 7479016
  15. Gene transfer into the medfly, Ceratitis capitata, with a Drosophila hydei transposable element.
    Science. 1995 Dec 22;270(5244):2002-5 PMID: 8533094
  16. Modifications of vectors pEF-BOS, pcDNA1 and pcDNA3 result in improved convenience and expression.
    Biotechniques. 1996 Dec;21(6):1013-5 PMID: 8969825
  17. Functional genomics in mice by tagged sequence mutagenesis.
    Nat Genet. 1997 Aug;16(4):338-44 PMID: 9241269
  18. Molecular reconstruction of Sleeping Beauty, a Tc1-like transposon from fish, and its transposition in human cells.
    Cell. 1997 Nov 14;91(4):501-10 PMID: 9390559
  19. Disruption and sequence identification of 2,000 genes in mouse embryonic stem cells.
    Nature. 1998 Apr 9;392(6676):608-11 PMID: 9560157
  20. A method for analyzing the qualitative and quantitative aspects of gene expression: a transcriptional profile revealed for HeLa cells.
    Nucleic Acids Res. 1998 Jun 1;26(11):2586-92 PMID: 9592141
  21. Transposon Tc1 of the nematode Caenorhabditis elegans jumps in human cells.
    Nucleic Acids Res. 1998 Jun 15;26(12):3013-7 PMID: 9611249
  22. The Himar1 mariner transposase cloned in a recombinant adenovirus vector is functional in mammalian cells.
    Nucleic Acids Res. 1998 Aug 15;26(16):3687-93 PMID: 9685483
  23. Chromosomal transposition of a Tc1/mariner-like element in mouse embryonic stem cells.
    Proc Natl Acad Sci U S A. 1998 Sep 1;95(18):10769-73 PMID: 9724779
  24. Efficient gene trap screening for novel developmental genes using IRES beta geo vector and in vitro preselection.
    Exp Cell Res. 1998 Oct 10;244(1):125-36 PMID: 9770356
  25. A genome-wide functional assay of signal transduction in living mammalian cells.
    Nat Biotechnol. 1998 Dec;16(13):1329-33 PMID: 9853613
  26. Trans-kingdom transposition of the Drosophila element mariner within the protozoan Leishmania.
    Science. 1997 Jun 13;276(5319):1716-9 PMID: 9180085
  27. Establishment of a gene-trap sequence tag library to generate mutant mice from embryonic stem cells.
    Nat Genet. 2000 Jan;24(1):13-4 PMID: 10615117
  28. Toward Anopheles transformation: Minos element activity in anopheline cells and embryos.
    Proc Natl Acad Sci U S A. 2000 Feb 29;97(5):2157-62 PMID: 10681436
  29. Saturation mutagenesis using maize transposons.
    Curr Opin Plant Biol. 2000 Apr;3(2):103-7 PMID: 10712955
  30. Functional genomics in Arabidopsis: large-scale insertional mutagenesis complements the genome sequencing project.
    Curr Opin Biotechnol. 2000 Apr;11(2):157-61 PMID: 10753770
  31. Mobility assays confirm the broad host-range activity of the Minos transposable element and validate new transformation tools.
    Insect Mol Biol. 2000 Jun;9(3):269-75 PMID: 10886410
Article Info
Journal
EMBO reports
Abbr.
EMBO Rep
ISSN
1469-221X
DOI
10.1093/embo-reports/kvd089
Published
2000-11-00
Pages
416-21
Language
English
Region
England
NLM ID
100963049
PMCID
PMC1083762
Subset
IM
External Links
PubMed source DOI: 10.1093/embo-reports/kvd089 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: [email protected]

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2