PUBLICATION

Reverse Genetic Screening Reveals Poor Correlation between Morpholino-Induced and Mutant Phenotypes in Zebrafish

Authors
Kok, F.O., Shin, M., Ni, C., Gupta, A., Grosse, A.S., van Impel, A., Kirchmaier, B.C., Peterson-Maduro, J., Kourkoulis, G., Male, I., DeSantis, D.F., Sheppard-Tindell, S., Ebarasi, L., Betsholtz, C., Schulte-Merker, S., Wolfe, S.A., Lawson, N.D.
ID
ZDB-PUB-141224-21
Date
2015
Source
Developmental Cell   32(1): 97-108 (Journal)
Registered Authors
Betsholtz, Christer, Ebarasi, Lwaki, Gupta, Ankit, Kourkoulis, George D., Lawson, Nathan, Peterson-Maduroe, Josi, Schulte-Merker, Stefan, Shin, Masahiro, van Impel, Andreas, Wolfe, Scot A.
Keywords
none
MeSH Terms
  • Zebrafish/genetics*
  • Zebrafish/growth & development
  • Gene Expression Regulation, Developmental
  • Reverse Transcriptase Polymerase Chain Reaction
  • Phenotype
  • Mutation/genetics*
  • RNA, Messenger/genetics
  • Animals
  • Deoxyribonucleases/genetics*
  • Deoxyribonucleases/metabolism
  • Blotting, Western
  • Zebrafish Proteins/antagonists & inhibitors
  • Zebrafish Proteins/genetics*
  • Oligonucleotides, Antisense/pharmacology*
  • Morpholinos/pharmacology*
  • Gene Expression Regulation/drug effects*
  • Gene Knockdown Techniques/methods*
  • Real-Time Polymerase Chain Reaction
(all 18)
PubMed
25533206 Full text @ Dev. Cell
Abstract
The widespread availability of programmable site-specific nucleases now enables targeted gene disruption in the zebrafish. In this study, we applied site-specific nucleases to generate zebrafish lines bearing individual mutations in more than 20 genes. We found that mutations in only a small proportion of genes caused defects in embryogenesis. Moreover, mutants for ten different genes failed to recapitulate published Morpholino-induced phenotypes (morphants). The absence of phenotypes in mutant embryos was not likely due to maternal effects or failure to eliminate gene function. Consistently, a comparison of published morphant defects with the Sanger Zebrafish Mutation Project revealed that approximately 80% of morphant phenotypes were not observed in mutant embryos, similar to our mutant collection. Based on these results, we suggest that mutant phenotypes become the standard metric to define gene function in zebrafish, after which Morpholinos that recapitulate respective phenotypes could be reliably applied for ancillary analyses.
Genes / Markers
Figures
Figure Gallery (6 images)
Show all Figures
Expression
Phenotype
Fish Conditions Stage Phenotype Figure
WT + MO3-birc6-as2standard conditionsLong-pec
ackr5um72/um72standard conditionsPrim-5 to Day 5
ackr5um73/um73standard conditionsPrim-5 to Day 5
amotum76/um76standard conditionsPrim-5 to Day 5
amotum77/um77standard conditionsPrim-5 to Day 5
angpt2aum94/um94standard conditionsPrim-5 to Day 5
arhgef9bum45/um45standard conditionsPrim-5 to Day 5
arhgef9bum45/um45standard conditionsDay 5
arhgef9bum45/um45standard conditionsDay 5
arhgef9bum45/um45standard conditionsDay 5
1 - 10 of 93
Show
Mutations / Transgenics
Allele Construct Type Affected Genomic Region
hu3072
    		Point Mutation
    hu3559
      		Point Mutation
      hu5333TgTransgenic Insertion
        hu7135TgTransgenic Insertion
          hu9963
            		Small Deletion
            hu10535
              		Small Deletion
              hu10965
                		Small Deletion
                s843TgTransgenic Insertion
                  sa19
                    		Point Mutation
                    sa56
                      		Point Mutation
                      1 - 10 of 62
                      Show
                      Human Disease / Model
                      No data available
                      Sequence Targeting Reagents
                      Target Reagent Reagent Type
                      birc6-as2MO3-birc6-as2MRPHLNO
                      birc6-as2TALEN1-birc6-as2TALEN
                      birc6-as2TALEN2-birc6-as2TALEN
                      ccbe1TALEN1-ccbe1TALEN
                      ets1CRISPR1-ets1CRISPR
                      flt4TALEN4-flt4TALEN
                      flt4TALEN5-flt4TALEN
                      fmnl3TALEN1-fmnl3TALEN
                      mmp2TALEN1-mmp2TALEN
                      nrp1aTALEN1-nrp1aTALEN
                      1 - 10 of 11
                      Show
                      Fish
                      Fish
                      ackr5um72/um72
                      ackr5um73/um73
                      amotum76/um76
                      amotum77/um77
                      angpt2aum94/um94
                      arhgef9bum45/um45
                      arhgef9bum45/um45; s843Tg
                      arhgef9bum45/um45; y1Tg
                      arrb1sa11194/sa11194
                      birc6-as2um158/um158
                      1 - 10 of 53
                      Show
                      Antibodies
                      Name Type Antigen Genes Isotypes Host Organism
                      Ab1-pdgfrbpolyclonalRabbit
                      1 - 1 of 1
                      Show
                      Orthology
                      No data available
                      Engineered Foreign Genes
                      Marker Marker Type Name
                      CitrineEFGCitrine
                      DsRedEFGDsRed
                      EGFPEFGEGFP
                      RFPEFGRFP
                      1 - 4 of 4
                      Show
                      Mapping
                      No data available
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                      Citation
                      Kok, F.O., Shin, M., Ni, C., Gupta, A., Grosse, A.S., van Impel, A., Kirchmaier, B.C., Peterson-Maduro, J., Kourkoulis, G., Male, I., DeSantis, D.F., Sheppard-Tindell, S., Ebarasi, L., Betsholtz, C., Schulte-Merker, S., Wolfe, S.A., Lawson, N.D. (2015) Reverse Genetic Screening Reveals Poor Correlation between Morpholino-Induced and Mutant Phenotypes in Zebrafish. Developmental Cell. 32(1):97-108.