A universal vector concept for a direct genotyping of transgenic organisms and a systematic creation of homozygous lines

  • Diploid transgenic organisms are either hemi- or homozygous. Genetic assays are, therefore, required to identify the genotype. Our AGameOfClones vector concept uses two clearly distinguishable transformation markers embedded in interweaved, but incompatible Lox site pairs. Cre-mediated recombination leads to hemizygous individuals that carry only one marker. In the following generation, heterozygous descendants are identified by the presence of both markers and produce homozygous progeny that are selected by the lack of one marker. We prove our concept in Tribolium castaneum by systematically creating multiple functional homozygous transgenic lines suitable for long-term fluorescence live imaging. Our approach saves resources and simplifies transgenic organism handling. Since the concept relies on the universal Cre-Lox system, it is expected to work in all diploid model organisms, for example, insects, zebrafish, rodents and plants. With appropriate adaptions, it can be used in knock-out assays to preselect homozygous individuals and thus minimize the number of wasted animals.
Metadaten
Author:Frederic Strobl, Anita Anderl, Ernst H. K. StelzerORCiDGND
URN:urn:nbn:de:hebis:30:3-463769
DOI:https://doi.org/10.7554/eLife.31677
ISSN:2050-084X
Pubmed Id:https://pubmed.ncbi.nlm.nih.gov/29543587
Parent Title (English):eLife
Publisher:eLife Sciences Publications
Place of publication:Cambridge
Contributor(s):Bruce Edgar
Document Type:Article
Language:English
Year of Completion:2018
Date of first Publication:2018/03/15
Publishing Institution:Universit├Ątsbibliothek Johann Christian Senckenberg
Release Date:2018/05/15
Tag:Chromosomes and gene expression; Cre-lox system; Developmental biology and stem cells; Genotyping; Light sheet-based fluorescence microscopy; Long-term live imaging; Tools and resources; Transgenesis; Tribolium castaneum
Volume:7
Issue:e31677
Page Number:21
First Page:1
Last Page:21
Note:
Copyright Strobl et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
HeBIS-PPN:433868910
Institutes:Biowissenschaften / Biowissenschaften
Exzellenzcluster / Exzellenzcluster Makromolekulare Komplexe
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Sammlungen:Universit├Ątspublikationen
Sammlung Biologie / Sondersammelgebiets-Volltexte
Open-Access-Publikationsfonds:Biowissenschaften
Licence (German):License LogoCreative Commons - Namensnennung 4.0