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• Consequently, we recommend injection into homozygous landing site embryos to provide optimal transgenesis efficiency.
• Similar integrations have been well-documented in mice when targeting the attP sites in the H11 locus, documenting background integrations as a universal caveat of vector-based transgene injections 18,21. Nonetheless, our genotyping PCR to verify proper transgene integration into attP landing sites together with careful screening enable selection o...
• Although both transgenesis methods achieved F0 embryos with 80-100% fluorescent coverage across notochords for the wild type Hs_I reporters (Fig. 4B,C,E), pIGLET-based injections yielded a significantly higher percentage of nearly complete notochord reporter activity among injected embryos (Tol2: 4.72% vs. pIGLET: 27.42%) (Fig. 4E)
• In addition to our previously established attB-containing pDEST backbone vectors (pCM268, no transgenesis marker; pCM327, cryaa:Venus) 22, we generated six additional backbone vectors for different cloning approaches: pRL055 pDESTattB_cryaa:Venus (inverted), pRL56 pDESTattB_exorh:EGFP, pCK122 pDESTattB_exorh:mCherry, pCK123 pDESTattB_exorh:mCerulea...

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