RoMo: An efficient strategy for functional mosaic analysis via stochastic Cre recombination and gene targeting in the ROSA26 locus

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RoMo: An efficient strategy for functional mosaic analysis via stochastic Cre recombination and gene targeting in the ROSA26 locus. / Movahedi, Kiavash; Wiegmann, Robert; De Vlaminck, Karen; Van Ginderachter, Jo A; Nikolaev, Viacheslav O.

In: BIOTECHNOL BIOENG, Vol. 115, No. 7, 07.2018, p. 1778-1792.

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@article{20c49593f9e74747b1062cb376d184b0,
title = "RoMo: An efficient strategy for functional mosaic analysis via stochastic Cre recombination and gene targeting in the ROSA26 locus",
abstract = "Functional mosaic analysis allows for the direct comparison of mutant cells with differentially marked control cells in the same organism. While this offers a powerful approach for elucidating the role of specific genes or signalling pathways in cell populations of interest, genetic strategies for generating functional mosaicism remain challenging. We describe a novel and streamlined approach for functional mosaic analysis, which combines stochastic Cre/lox recombination with gene targeting in the ROSA26 locus. With the RoMo strategy a cell population of interest is randomly split into a cyan fluorescent and red fluorescent subset, of which the latter overexpresses a chosen transgene. To integrate this approach into high-throughput gene targeting initiatives, we developed a procedure that utilizes Gateway cloning for the generation of new targeting vectors. RoMo can be used for gain-of-function experiments or for altering signaling pathways in a mosaic fashion. To demonstrate this, we developed RoMo-dnGs mice, in which Cre-recombined red fluorescent cells co-express a dominant-negative Gs protein. RoMo-dnGs mice allowed us to inhibit G protein-coupled receptor activation in a fraction of cells, which could then be directly compared to differentially marked control cells in the same animal. We demonstrate how RoMo-dnGs mice can be used to obtain mosaicism in the brain and in peripheral organs for various cell types. RoMo offers an efficient new approach for functional mosaic analysis that extends the current toolbox and may reveal important new insights into in vivo gene function.",
keywords = "Journal Article",
author = "Kiavash Movahedi and Robert Wiegmann and {De Vlaminck}, Karen and {Van Ginderachter}, {Jo A} and Nikolaev, {Viacheslav O}",
note = "{\textcopyright} 2018 Wiley Periodicals, Inc.",
year = "2018",
month = jul,
doi = "10.1002/bit.26594",
language = "English",
volume = "115",
pages = "1778--1792",
journal = "BIOTECHNOL BIOENG",
issn = "0006-3592",
publisher = "Wiley-VCH Verlag GmbH",
number = "7",

}

RIS

TY - JOUR

T1 - RoMo: An efficient strategy for functional mosaic analysis via stochastic Cre recombination and gene targeting in the ROSA26 locus

AU - Movahedi, Kiavash

AU - Wiegmann, Robert

AU - De Vlaminck, Karen

AU - Van Ginderachter, Jo A

AU - Nikolaev, Viacheslav O

N1 - © 2018 Wiley Periodicals, Inc.

PY - 2018/7

Y1 - 2018/7

N2 - Functional mosaic analysis allows for the direct comparison of mutant cells with differentially marked control cells in the same organism. While this offers a powerful approach for elucidating the role of specific genes or signalling pathways in cell populations of interest, genetic strategies for generating functional mosaicism remain challenging. We describe a novel and streamlined approach for functional mosaic analysis, which combines stochastic Cre/lox recombination with gene targeting in the ROSA26 locus. With the RoMo strategy a cell population of interest is randomly split into a cyan fluorescent and red fluorescent subset, of which the latter overexpresses a chosen transgene. To integrate this approach into high-throughput gene targeting initiatives, we developed a procedure that utilizes Gateway cloning for the generation of new targeting vectors. RoMo can be used for gain-of-function experiments or for altering signaling pathways in a mosaic fashion. To demonstrate this, we developed RoMo-dnGs mice, in which Cre-recombined red fluorescent cells co-express a dominant-negative Gs protein. RoMo-dnGs mice allowed us to inhibit G protein-coupled receptor activation in a fraction of cells, which could then be directly compared to differentially marked control cells in the same animal. We demonstrate how RoMo-dnGs mice can be used to obtain mosaicism in the brain and in peripheral organs for various cell types. RoMo offers an efficient new approach for functional mosaic analysis that extends the current toolbox and may reveal important new insights into in vivo gene function.

AB - Functional mosaic analysis allows for the direct comparison of mutant cells with differentially marked control cells in the same organism. While this offers a powerful approach for elucidating the role of specific genes or signalling pathways in cell populations of interest, genetic strategies for generating functional mosaicism remain challenging. We describe a novel and streamlined approach for functional mosaic analysis, which combines stochastic Cre/lox recombination with gene targeting in the ROSA26 locus. With the RoMo strategy a cell population of interest is randomly split into a cyan fluorescent and red fluorescent subset, of which the latter overexpresses a chosen transgene. To integrate this approach into high-throughput gene targeting initiatives, we developed a procedure that utilizes Gateway cloning for the generation of new targeting vectors. RoMo can be used for gain-of-function experiments or for altering signaling pathways in a mosaic fashion. To demonstrate this, we developed RoMo-dnGs mice, in which Cre-recombined red fluorescent cells co-express a dominant-negative Gs protein. RoMo-dnGs mice allowed us to inhibit G protein-coupled receptor activation in a fraction of cells, which could then be directly compared to differentially marked control cells in the same animal. We demonstrate how RoMo-dnGs mice can be used to obtain mosaicism in the brain and in peripheral organs for various cell types. RoMo offers an efficient new approach for functional mosaic analysis that extends the current toolbox and may reveal important new insights into in vivo gene function.

KW - Journal Article

U2 - 10.1002/bit.26594

DO - 10.1002/bit.26594

M3 - SCORING: Journal article

C2 - 29573361

VL - 115

SP - 1778

EP - 1792

JO - BIOTECHNOL BIOENG

JF - BIOTECHNOL BIOENG

SN - 0006-3592

IS - 7

ER -