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Methods Mol Biol. 2020;2092:109-122. doi: 10.1007/978-1-0716-0175-4_9.

A Simple Guide for Generating BAC Transgenic Animals for Retinal Research.

Agca C1,2,3, Grimm C4,5,6.

Author information

1
Lab for Retinal Cell Biology, Department of Ophthalmology, University of Zurich, Zurich, Switzerland. cavit.agca@sabanciuniv.edu.
2
Molecular Biology, Genetics and Bioengineering Program, Sabanci University, Istanbul, Turkey. cavit.agca@sabanciuniv.edu.
3
Nanotechnology Research and Application Center (SUNUM), Sabanci University, Istanbul, Turkey. cavit.agca@sabanciuniv.edu.
4
Lab for Retinal Cell Biology, Department of Ophthalmology, University of Zurich, Zurich, Switzerland.
5
Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Zurich, Switzerland.
6
Neuroscience Center (ZNZ), University of Zurich, Zurich, Switzerland.

Abstract

Bacterial artificial chromosomes (BACs) are genomic tools that can carry several hundred kilobases of exogenous genomic material. This allows to incorporate sufficiently large DNA stretches to include most if not all upstream and downstream cis-regulatory elements of a gene in order to mimic and analyze its endogenous regulation of expression using a reporter protein in vivo. Here, we illustrate the generation of a BAC:LIF-EGFP transgenic mouse line to describe a simplified version of BAC transgenesis using galK-based recombineering.

KEYWORDS:

BAC sequencing; BAC transgenics; Colony real-time PCR; Electroporation; Lif; Minimal media; Recombineering; SW102; galK

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