Microinjection of Zebrafish Embryos

Zebrafish embryos were injected using glass capillary needles (30-0020, Harvard Apparatus, MA, USA), which were pulled by a needle puller (P-97, Sutter Instrument) and attached to a microinjector system (PV820, World Precision Instruments). Microinjections of mRNAs and morpholinos (MOs) were performed at the one-cell stage. mRNAs were synthesized using mMACHINE SP6 kit (Ambion). The following mRNAs were injected: 70 pg membrane RFP (Iioka et al., 2004 (link)), 70 pg H2B GFP (Keller et al., 2008 (link)), 200 pg CAMypt1 (Schwayer et al., 2019 (link)), and 72 pg chk1 (Shamipour et al., 2019 (link)). The following MOs were injected: 4 ng e-cadherin MO, 4 ng control MO for e-cadherin (Babb and Marrs, 2004 (link)). To induce mesodermal progenitor cell fate, the following combination of mRNAs and MOs was injected: 100 pg ndr2l cyclops mRNA and 2 ng casanova MO (Krieg et al., 2008 (link)). To label the interstitial fluid, 1 nL of 0.6 mg/ml dextran Alexa Fluor 647 (10,000 MW; D22914, Invitrogen) was injected in the blastoderm of 1k-stage embryos (∼3 hpf).

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Petridou N.I., Corominas-Murtra B., Heisenberg C.P, & Hannezo E. (2021). Rigidity percolation uncovers a structural basis for embryonic tissue phase transitions. Cell, 184(7), 1914-1928.e19.

Publication 2021

PV820 mMACHINE SP6 kit

Alexa fluor 647 Blastoderm Cadherin Cadherin 4 Capillary Cell Cyclops Dextran Embryos Interstitial fluid Membrane Mesodermal Microinjections Morpholinos Mrna Needle Progenitor cell Zebrafish

Corresponding Organization : Institute of Science and Technology Austria

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Variable analysis

independent variables

Injection of mRNAs: 70 pg membrane RFP, 70 pg H2B GFP, 200 pg CAMypt1, and 72 pg chk1
Injection of morpholinos (MOs): 4 ng e-cadherin MO, 4 ng control MO for e-cadherin
Injection of 100 pg ndr2l cyclops mRNA and 2 ng casanova MO to induce mesodermal progenitor cell fate
Injection of 1 nL of 0.6 mg/ml dextran Alexa Fluor 647 (10,000 MW) in the blastoderm of 1k-stage embryos (∼3 hpf) to label the interstitial fluid

dependent variables

Not explicitly mentioned

control variables

Glass capillary needles (30-0020, Harvard Apparatus, MA, USA)
Needle puller (P-97, Sutter Instrument)
Microinjector system (PV820, World Precision Instruments)
Microinjections performed at the one-cell stage
MRNAs synthesized using mMACHINE SP6 kit (Ambion)

positive controls

4 ng control MO for e-cadherin

negative controls

None specified

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