Biology Reference
In-Depth Information
CHAPTER 3
Genome Engineering by Transgene-
Instructed Gene Conversion in C. elegans
Valerie J.P. Robert
*
,
y
,
z
,
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and Jean-Louis Bessereau
*
,
y
,
z
*
Ecole Normale Superieure, Institut de Biologie de l
'
ENS, IBENS, Paris, France
y
INSERM U1024, Paris, France
z
CNRS, UMR 8197, Paris, France
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Present address: Laboratory of Molecular and Cellular Biology, UMR5239 CNRS, Ecole Normale
Superieure de Lyon, Lyon, France
Abstract
I. Introduction
A. Transgenesis in C. elegans
B. Genome Engineering by Homologous Recombination in C. elegans
II. General Features of the MosTIC Technique and Experimental Outline
III. Methods
A. Identification of a Mos1 Insertion in the Target Region
B. Construction of Transgenic Lines
C. Heat-Shock Induction of Mos1 Excision and Pooling of Heat-Shocked Animals
D. Screening
E. An Alternative Protocol Based on the Constitutive Expression of the Mos
Transposase
F. An Alternative Protocol for MosTIC-Engineered KO/del Alleles Selection
G. Frequently Asked Questions
IV. Materials
A. Equipment
B. Solutions and Reagents
C. Plasmids
V. Discussion: MosTIC Applications and Derivatives
A. Gene Function Analysis Using MosTIC-Engineered Alleles
B. Single-Copy Transgene Genomic Integration by MosSCI
VI. Summary
Acknowledgment
References
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