PUBLICATION

Enhancer trapping in zebrafish using the Sleeping Beauty transposon

Authors
Balciunas, D., Davidson, A.E., Sivasubbu, S., Hermanson, S.B., Welle, Z., and Ekker, S.C.
ID
ZDB-PUB-040908-16
Date
2004
Source
BMC Genomics   5(1): 62 (Journal)
Registered Authors
Balciunas, Darius, Ekker, Stephen C., Hermanson, Spencer, Sivasubbu, Sridhar
Keywords
none
MeSH Terms
  • Sequence Analysis, DNA/methods
  • Enhancer Elements, Genetic*
  • Animals
  • Motor Neurons/metabolism
  • Promoter Regions, Genetic
  • Zebrafish/embryology
  • Zebrafish/genetics*
  • Zebrafish/metabolism
  • Green Fluorescent Proteins/biosynthesis
  • DNA Transposable Elements*
  • Genomics/methods*
  • Embryonic Development/genetics
  • Germ Cells
  • In Situ Hybridization
  • Polymerase Chain Reaction
  • Gene Transfer Techniques
  • Pilot Projects
  • Glycoside Hydrolases/genetics
(all 18)
PubMed
15347431 Full text @ BMC Genomics
Abstract
BACKGROUND: Among functional elements of a metazoan gene, enhancers are particularly difficult to find and annotate. Pioneering experiments in Drosophila have demonstrated the value of enhancer "trapping" using an invertebrate to address this functional genomics problem. RESULTS: We modulated a Sleeping Beauty transposon-based transgenesis cassette to establish an enhancer trapping technique for use in a vertebrate model system, zebrafish Danio rerio. We established 9 lines of zebrafish with distinct tissue- or organ- specific GFP expression patterns from 90 founders that produced GFP-expressing progeny. We have molecularly characterized these lines and show that in each line, a specific GFP expression pattern is due to a single transposition event. Many of the insertions are into introns of zebrafish genes predicted in the current genome assembly. We have identified both previously characterized as well as novel expression patterns from this pilot screen. For example, the ET7 line harbors a transposon insertion near the mkp3 locus and expresses GFP in the midbrain-hindbrain boundary, forebrain and the ventricle, matching a subset of the known FGF8-dependendent mkp3 expression domain. The ET2 line, in contrast, expresses GFP specifically in caudal primary motoneurons due to an insertion into the poly(ADP-ribose) glycohydrolase (PARG) locus. This surprising expression pattern was confirmed using in situ hybridization techniques for the endogenous PARG mRNA, indicating the enhancer trap has replicated this unexpected and highly localized PARG expression with good fidelity. Finally, we show that it is possible to excise a Sleeping Beauty transposon from a genomic location in the zebrafish germline. CONCLUSIONS: This genomics tool offers the opportunity for large-scale biological approaches combining both expression and genomic-level sequence analysis using as a template an entire vertebrate genome.
Genes / Markers
Figures
Figure Gallery (5 images)
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Expression
Gene Antibody Fish Conditions Stage Qualifier Anatomy Assay Figure
dusp6WTstandard conditions26+ somitesISH
GFPmbipmn3Etstandard conditionsPrim-25IFL
GFPmbipmn3Etstandard conditions5-9 somitesIFL
GFPmn1Etstandard conditions20-25 somites to Prim-25IFL
GFPmn1Etstandard conditions5-9 somitesIFL
GFPmn4Etstandard conditionsDay 5Not DetectedIFL
GFPmn4Etstandard conditions14-19 somites to High-pecIFL
GFPmn5Etstandard conditionsPrim-5IFL
GFPmn6Etstandard conditionsProtruding-mouthNot DetectedIFL
GFPmn6Etstandard conditionsPrim-5IFL
1 - 10 of 30
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Phenotype
No data available
Mutations / Transgenics
Allele Construct Type Affected Genomic Region
etl1_unspecified
    		Unspecified
    etl4_unspecified
      		Unspecified
      etl5_unspecified
        		Unspecified
        etl6_unspecified
          		Unspecified
          etl7_unspecified
            		Unspecified
            etl8_unspecified
              		Unspecified
              etl9_unspecified
                		Unspecified
                mbip_unspecified
                  		Unspecified
                  mn1EtTransgenic Insertion
                    mn2EtTransgenic Insertion
                    1 - 10 of 18
                    Show
                    Human Disease / Model
                    No data available
                    Sequence Targeting Reagents
                    No data available
                    Fish
                    Fish
                    mbipmn3Et
                    mn1Et
                    mn4Et
                    mn5Et
                    mn6Et
                    mn7Et
                    mn8Et
                    mn9Et
                    pargamn2Et
                    WT
                    1 - 10 of 10
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                    Antibodies
                    No data available
                    Orthology
                    No data available
                    Engineered Foreign Genes
                    Marker Marker Type Name
                    GFPEFGGFP
                    1 - 1 of 1
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                    Mapping
                    Entity Type Entity Symbol Location
                    Featuremn7EtUnmapped
                    GENEdusp6Chr: 25 Details
                    1 - 2 of 2
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                    Citation
                    Balciunas, D., Davidson, A.E., Sivasubbu, S., Hermanson, S.B., Welle, Z., and Ekker, S.C. (2004) Enhancer trapping in zebrafish using the Sleeping Beauty transposon. BMC Genomics. 5(1):62.