Green fluorescent protein tagging Drosophila proteins at their native genomic loci with small P elements

Peter J Clyne; Jennie S Brotman; Sean T Sweeney; Graeme Davis

Green fluorescent protein tagging Drosophila proteins at their native genomic loci with small P elements

Peter J Clyne, Jennie S Brotman, Sean T Sweeney, Graeme Davis

Biology

Research output: Contribution to journal › Article › peer-review

Abstract

We describe a technique to tag Drosophila proteins with GFP at their native genomic loci. This technique uses a new, small P transposable element (the Wee-P) that is composed primarily of the green fluorescent protein (GFP) sequence flanked by consensus splice acceptor and splice donor sequences. We demonstrate that insertion of the Wee-P can generate GFP fusions with native proteins. We further demonstrate that GFP-tagged proteins have correct subcellular localization and can be expressed at near-normal levels. We have used the Wee-P to tag genes with a wide variety of functions, including transmembrane proteins. A genetic analysis of 12 representative fusion lines demonstrates that loss-of-function phenotypes are not caused by the Wee-P insertion. This technology allows the generation of GFP-tagged reagents on a genome-wide scale with diverse potential applications.

Original language	English
Pages (from-to)	1433-41
Number of pages	9
Journal	Genetics
Volume	165
Issue number	3
Publication status	Published - 2003

Keywords

Animals
Artificial Gene Fusion
Base Sequence
DNA Primers
DNA Transposable Elements
Drosophila
Drosophila Proteins
Genomics
Green Fluorescent Proteins
Luminescent Proteins
Recombinant Fusion Proteins
Subcellular Fractions

Cite this

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title = "Green fluorescent protein tagging Drosophila proteins at their native genomic loci with small P elements",

abstract = "We describe a technique to tag Drosophila proteins with GFP at their native genomic loci. This technique uses a new, small P transposable element (the Wee-P) that is composed primarily of the green fluorescent protein (GFP) sequence flanked by consensus splice acceptor and splice donor sequences. We demonstrate that insertion of the Wee-P can generate GFP fusions with native proteins. We further demonstrate that GFP-tagged proteins have correct subcellular localization and can be expressed at near-normal levels. We have used the Wee-P to tag genes with a wide variety of functions, including transmembrane proteins. A genetic analysis of 12 representative fusion lines demonstrates that loss-of-function phenotypes are not caused by the Wee-P insertion. This technology allows the generation of GFP-tagged reagents on a genome-wide scale with diverse potential applications.",

keywords = "Animals, Artificial Gene Fusion, Base Sequence, DNA Primers, DNA Transposable Elements, Drosophila, Drosophila Proteins, Genomics, Green Fluorescent Proteins, Luminescent Proteins, Recombinant Fusion Proteins, Subcellular Fractions",

author = "Clyne, {Peter J} and Brotman, {Jennie S} and Sweeney, {Sean T} and Graeme Davis",

year = "2003",

language = "English",

volume = "165",

pages = "1433--41",

journal = "Genetics",

issn = "0016-6731",

publisher = "Genetics Society of America",

number = "3",

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TY - JOUR

T1 - Green fluorescent protein tagging Drosophila proteins at their native genomic loci with small P elements

AU - Clyne, Peter J

AU - Brotman, Jennie S

AU - Sweeney, Sean T

AU - Davis, Graeme

PY - 2003

Y1 - 2003

N2 - We describe a technique to tag Drosophila proteins with GFP at their native genomic loci. This technique uses a new, small P transposable element (the Wee-P) that is composed primarily of the green fluorescent protein (GFP) sequence flanked by consensus splice acceptor and splice donor sequences. We demonstrate that insertion of the Wee-P can generate GFP fusions with native proteins. We further demonstrate that GFP-tagged proteins have correct subcellular localization and can be expressed at near-normal levels. We have used the Wee-P to tag genes with a wide variety of functions, including transmembrane proteins. A genetic analysis of 12 representative fusion lines demonstrates that loss-of-function phenotypes are not caused by the Wee-P insertion. This technology allows the generation of GFP-tagged reagents on a genome-wide scale with diverse potential applications.

AB - We describe a technique to tag Drosophila proteins with GFP at their native genomic loci. This technique uses a new, small P transposable element (the Wee-P) that is composed primarily of the green fluorescent protein (GFP) sequence flanked by consensus splice acceptor and splice donor sequences. We demonstrate that insertion of the Wee-P can generate GFP fusions with native proteins. We further demonstrate that GFP-tagged proteins have correct subcellular localization and can be expressed at near-normal levels. We have used the Wee-P to tag genes with a wide variety of functions, including transmembrane proteins. A genetic analysis of 12 representative fusion lines demonstrates that loss-of-function phenotypes are not caused by the Wee-P insertion. This technology allows the generation of GFP-tagged reagents on a genome-wide scale with diverse potential applications.

KW - Animals

KW - Artificial Gene Fusion

KW - Base Sequence

KW - DNA Primers

KW - DNA Transposable Elements

KW - Drosophila

KW - Drosophila Proteins

KW - Genomics

KW - Green Fluorescent Proteins

KW - Luminescent Proteins

KW - Recombinant Fusion Proteins

KW - Subcellular Fractions

M3 - Article

C2 - 14668392

SN - 0016-6731

VL - 165

SP - 1433

EP - 1441

JO - Genetics

JF - Genetics

IS - 3

ER -