Microinjection was performed as previously described (5 (link)). Two gene-specific MO-substituted antisense oligonucleotides (5′-AAAATATATGCATTCATGTTGATCA-3′) complementary to the start codon region of the sea urchin Cara7 gene and a second MO (5′-CGACCTTCAGATATATTCTCACAAA-3′) complementary to the 5′-untranslated region were obtained from Gene Tools. In addition, a scramble MO 5′-CCTCTTACCTCAGTTACAATTTATA-3′ that has no biological target in the sea urchin was used to obtain the control groups. The MOs were dissolved in injection Buffer (20 mM Hepes, 120 mM KCl, 24% glycerol, pH 8.0) and were injected into the freshly fertilized egg (one-cell stage) using a microinjection system (Picospritzer III, Parker) mounted on an inverse microscope (Zeiss Observer D1) equipped with a cooling stage.
Protocol detail
Microinjection of Antisense Oligonucleotides in Sea Urchin
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Antisense oligonucleotides
Biological
Buffer
Cell
Fertilized egg
Gene
Glycerol
Hepes
Microinjection
Microscope
Sea urchin
Start codon
Untranslated region
Corresponding Organization :
Other organizations : Kiel University
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Variable analysis
independent variables
- Two gene-specific MO-substituted antisense oligonucleotides (5′-AAAATATATGCATTCATGTTGATCA-3′) complementary to the start codon region of the sea urchin Cara7 gene
- A second MO (5′-CGACCTTCAGATATATTCTCACAAA-3′) complementary to the 5′-untranslated region
- Not explicitly mentioned
- A scramble MO 5′-CCTCTTACCTCAGTTACAATTTATA-3′ that has no biological target in the sea urchin
- Positive control: Not specified
- Negative control: Scramble MO 5′-CCTCTTACCTCAGTTACAATTTATA-3′ that has no biological target in the sea urchin
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