A library of MiMICs allows tagging of genes and reversible, spatial and temporal knockdown of proteins in Drosophila

Author:

Nagarkar-Jaiswal Sonal1,Lee Pei-Tseng1,Campbell Megan E1,Chen Kuchuan2,Anguiano-Zarate Stephanie1,Cantu Gutierrez Manuel1,Busby Theodore1,Lin Wen-Wen1,He Yuchun3,Schulze Karen L3,Booth Benjamin W4,Evans-Holm Martha4,Venken Koen JT5,Levis Robert W6,Spradling Allan C6,Hoskins Roger A4,Bellen Hugo J12378

Affiliation:

1. Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, United States

2. Program in Developmental Biology, Baylor College of Medicine, Houston, United States

3. Howard Hughes Medical Institute, Baylor College of Medicine, Houston, United States

4. Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, United States

5. Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, United States

6. Department of Embryology, Howard Hughes Medical Institute, Carnegie Institution for Science, Baltimore, United States

7. Jan and Dan Duncan Neurological Research Institute, Texas Children's Hospital, Houston, United States

8. Department of Neuroscience, Baylor College of Medicine, Houston, United States

Abstract

Here, we document a collection of ∼7434 MiMIC (Minos Mediated Integration Cassette) insertions of which 2854 are inserted in coding introns. They allowed us to create a library of 400 GFP-tagged genes. We show that 72% of internally tagged proteins are functional, and that more than 90% can be imaged in unfixed tissues. Moreover, the tagged mRNAs can be knocked down by RNAi against GFP (iGFPi), and the tagged proteins can be efficiently knocked down by deGradFP technology. The phenotypes associated with RNA and protein knockdown typically correspond to severe loss of function or null mutant phenotypes. Finally, we demonstrate reversible, spatial, and temporal knockdown of tagged proteins in larvae and adult flies. This new strategy and collection of strains allows unprecedented in vivo manipulations in flies for many genes. These strategies will likely extend to vertebrates.

Funder

National Institute of General Medical Sciences (NIGMS)

National Institute of Child Health and Human Development (NICHD)

March of Dimes Foundation

Cancer Prevention and Research Institute of Texas (CPRIT)

National Human Genome Research Institute (NHGRI)

Baylor College of Medicine

National Institutes of Health (NIH)

Publisher

eLife Sciences Publications, Ltd

Subject

General Immunology and Microbiology,General Biochemistry, Genetics and Molecular Biology,General Medicine,General Neuroscience

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