Genetic Dissection of Heart Development
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cific in situ and antibody probes, in combination with the classic visual
analysis, searching for organogenesis mutations. We discovered mutations specifically affecting heart formation and/or function, organ laterality, the vascular system or the development of kidney, and the gut and
pancreas (Chen and Fishman, unpublished data). Detail characterization
and molecular cloning of these mutations is ongoing.
7.6 Cloning Zebrafish Mutations
Genetic screens provide means to dissect genetic pathways. The cloning
of these mutations leads to molecular understanding of the pathways. A
small fraction of the zebrafish mutations are generated by retrovirusbased insertion mutagenesis (Gaiano et al. 1996). The advantage of this
approach is that the mutation is "tagged" by the retrovirus and is easy to
clone. Once a desirable mutant phenotype is identified, the molecular
nature of the mutated gene can easily be revealed. However, the efficiency of the retrovirus-based mutagenesis is still significantly lower
than chemical mutagenesis and is labor-intensive (Schier et al. 1996).
Most of the zebrafish mutations currently available are chemicallyinduced point mutations. Although it is easier to generate mutations
with this approach, it is much more difficult to clone them. Up to now,
only two zebrafish mutations have been reported to be cloned positionally (Brownlie et al. 1998; Zhang et al. 1998). However, zebrafish
genomic resources have grown rapidly in the past few years, facilitating
positional cloning of the mutations. The first step toward cloning is to
place these mutations on a genetic map. Currently, about 2400 markers
are placed on the SSLP-based genetic map, giving the resolution of
0.9 cM on average (Shimoda et al. 1999). This resolution allows one to
initiate physical mapping and walking. Many mutations have been successfully placed on the SSLP map.
The most expeditious cloning strategy is through candidate genes.
About 20 zebrafish mutations have been cloned using as candidates,
genes identified because of their expression pattern and/or map position
(for examples see Hild et al. 1999; Kishimoto et al. 1997; Rauch et al.
1997). Mapping cDNA or EST by polymorphism (Gates et al. 1999) can
be quite time-consuming, but is rapid when using radiation hybrid
panels. Two zebrafish radiation hybrid panels are now available (Geisler
117
cific in situ and antibody probes, in combination with the classic visual
analysis, searching for organogenesis mutations. We discovered mutations specifically affecting heart formation and/or function, organ laterality, the vascular system or the development of kidney, and the gut and
pancreas (Chen and Fishman, unpublished data). Detail characterization
and molecular cloning of these mutations is ongoing.
7.6 Cloning Zebrafish Mutations
Genetic screens provide means to dissect genetic pathways. The cloning
of these mutations leads to molecular understanding of the pathways. A
small fraction of the zebrafish mutations are generated by retrovirusbased insertion mutagenesis (Gaiano et al. 1996). The advantage of this
approach is that the mutation is "tagged" by the retrovirus and is easy to
clone. Once a desirable mutant phenotype is identified, the molecular
nature of the mutated gene can easily be revealed. However, the efficiency of the retrovirus-based mutagenesis is still significantly lower
than chemical mutagenesis and is labor-intensive (Schier et al. 1996).
Most of the zebrafish mutations currently available are chemicallyinduced point mutations. Although it is easier to generate mutations
with this approach, it is much more difficult to clone them. Up to now,
only two zebrafish mutations have been reported to be cloned positionally (Brownlie et al. 1998; Zhang et al. 1998). However, zebrafish
genomic resources have grown rapidly in the past few years, facilitating
positional cloning of the mutations. The first step toward cloning is to
place these mutations on a genetic map. Currently, about 2400 markers
are placed on the SSLP-based genetic map, giving the resolution of
0.9 cM on average (Shimoda et al. 1999). This resolution allows one to
initiate physical mapping and walking. Many mutations have been successfully placed on the SSLP map.
The most expeditious cloning strategy is through candidate genes.
About 20 zebrafish mutations have been cloned using as candidates,
genes identified because of their expression pattern and/or map position
(for examples see Hild et al. 1999; Kishimoto et al. 1997; Rauch et al.
1997). Mapping cDNA or EST by polymorphism (Gates et al. 1999) can
be quite time-consuming, but is rapid when using radiation hybrid
panels. Two zebrafish radiation hybrid panels are now available (Geisler