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The art and design of genetic screens: zebrafish

Nature Reviews Genetics volume 2, pages 956–966 (2001)Cite this article

Key Points

  • The zebrafish offers a forward genetic system in which to explore vertebrate biological processes.

  • Zebrafish were used to carry out the first large-scale genetic screens in vertebrates, which identified more than 2,000 mutations involved in embryonic development.

  • Haploid and homozygous diploid screens expose recessive alleles in a generation, eliminating the need to work up an entire generation of fish.

  • Screens can use various mutagens, which offer different advantages in mutation rate or the ability to clone the gene.

  • Fluorescent reporter genes are used to visualize biological processes in zebrafish and have been used successfully in genetic screens to identify mutants (for example, digestive and retinal axon mutants).

  • Specific zebrafish behaviours are amenable to genetic screens, including vision response, locomotion and addiction behaviours.

  • Gene function can be explored in zebrafish by identifying mutations that enhance or suppress a phenotype, by isolating new and conditional alleles, as well as by using gene knockdown and gain-of-function technologies.

  • Small molecules can penetrate the chorion of zebrafish embryos and induce embryonic phenotypes within a controlled time during development.

  • Screen design requires a balance between selecting parameters that will best identify mutations of interest with those that are achievable in individual laboratories.

Abstract

Inventive genetic screens in zebrafish are revealing new genetic pathways that control vertebrate development, disease and behaviour. By exploiting the versatility of zebrafish, biological processes that had been previously obscured can be visualized and many of the responsible genes can be isolated. Coupled with gene knockdown and overexpression technologies, and small-molecule-induced phenotypes, genetic screens in zebrafish provide a powerful system by which to dissect vertebrate gene function and gene networks.

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Figure 1: Outline of large-scale F2 genetic screens.
Figure 2: Examples of mutants identified in zebrafish large-scale screening efforts.
Figure 3: Outline of insertional mutagenesis screen.
Figure 4: Fluorescent reporter screens.
Figure 5: The space cadet locomotion mutant.
Figure 6: Behaviour screens: visual adaptation mutants.
Figure 7: Morpholino-induced antisense phenotypes.

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Acknowledgements

We thank members of the Zon laboratory for helpful discussions and critical reading of the manuscript. E.E.P. is funded by a long-term postdoctoral fellowship from the Human Frontier Science Program. L.I.Z. is funded by the Howard Hughes Medical Institute and by a National Institutes of Health grant.

Author information

Authors and Affiliations

  1. Howard Hughes Medical Institute, Children's Hospital of Boston, 300 Longwood Avenue, Enders 750, Boston, 02115, Massachusetts, USA

    E. Elizabeth Patton & Leonard I. Zon

Authors
  1. E. Elizabeth Patton
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  2. Leonard I. Zon
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Corresponding author

Correspondence to Leonard I. Zon.

Related links

Related links

DATABASES

LocusLink 

CK

factor VII

Ferrochelatase

Ferroportin 1

fibrinogen

gata1

Hh

Mafb

mck

Nodal

shh

twhh

UDP-glucose dehydrogenase

UROD 

Medscape Drug Info 

atorvastatin 

OMIM 

erythropoietic protoporphyria

hepatoerythropoietic porphyria 

ZFIN 

ast

blw

bon

cas

cfs

cdy

clo

cyc

des

drc

fau

fss

hag

has

jek

krox20

lak

leo

lof

mic

nbb

nev

oep

pie

plx

reg

reg6

slj

smo

spc

sqt

sty

syu

unp

val

weh

yot

yng

yqe

FURTHER INFORMATION

Encyclopedia of Life Sciences: Zebrafish as an experimental organism

Zebrafish anatomy guide

ZFIN (Zebrafish Information Network)

Zon lab

Glossary

GYNOGENESIS

Development of an organism derived from the genetic material of the female gamete.

PARTHENOGENESIS

Development of an organism derived from an unfertilized gamete.

RHOMBOMERE

Each of seven neuroepithelial segments found in the embryonic hindbrain that adopt distinct molecular and cellular properties, restrictions in cell mixing and ordered domains of gene expression.

CHIASMA INTERFERENCE

The inhibition of crossover events during meiosis such that there is generally only one crossover event per chromosome arm.

SWIM BLADDER

An internal fish organ filled with gases, which are regulated to allow the fish to rise and fall. In some teleosts, the swim bladder can have a role in respiration and sound production and reception.

SOMITE

Paired cubical paraxial mesodermal segment, which is often used as a staging index during embryogenesis.

INVERSE PCR

A technique for amplifying DNA by PCR that uses primers that initiate replication in opposite directions to each other, as compared with standard PCR, which uses primers that initiate replication towards one another.

TECTUM

The dorsal portion of the midbrain (mesencephalon) that mediates reflexive responses to visual and auditory stimuli.

DOPAMINERGIC INTERPLEXIFORM CELL

(DI-IPC). Residing in the inner nuclear layer of the retina, this type of cell releases dopamine to regulate light adaptation in the retina.

TELEOST

Ray-finned bony fishes.

TURBIDOMETRY

A way to measure the solution turbidity, this can be used to assay for the formation of fibrin in the form of visible clotting in plasma.

HOLOPROSENCEPHALY

Failure of the forebrain (prosencephalon) to divide into hemispheres or lobes, often accompanied by a deficit in midline facial development.

RNA CAGING

RNA inactivation through the covalent attachment of a photo-removable synthetic compound called the caging group. RNA is reactivated by photo-illumination with a specific light wavelength.

GAL4/UAS SYSTEM

A genetic system for controlling the induction of gene expression. An activator line that expresses the yeast transcriptional activator GAL4 gene under the control of the heat-shock 70 promoter (hsp70) or a tissue-specific promoter is crossed to an effector line that carries the DNA-binding motif of Gal4 (UAS) fused to the gene of interest. As a result, the progeny of this cross expresses the gene of interest in an activator-specific manner.

CHORION

An extraembryonic membrane that surrounds the zebrafish embryo during the first 2 days of development.

OTOLITH

One of the small particles of calcium carbonate in the sacculus of the inner ear. Pressure of the otoliths on the hair cells of the macula (the most sensitive area of the ear) provide sensory inputs about acceleration and gravity.

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Patton, E., Zon, L. The art and design of genetic screens: zebrafish. Nat Rev Genet 2, 956–966 (2001). https://doi.org/10.1038/35103567

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