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Homology between the human LRRK2 and the zebrafish Lrrk2 proteins and generation of the zebrafish <italic toggle='yes'>lrrk2</italic><sup><italic toggle='yes'>tud113</italic></sup> allele.(A) Alignment of the whole sequence and the individual domains of human LRRK2 (NP_940980) and zebrafish Lrrk2 (NP_001188385) proteins revealed a high degree of conservation of the catalytic core. The percentages of identity (same residues at the same positions in the alignment) and similarity (identical residues plus conservative substitutions) are indicated. Abbreviations: aa, amino acids; ANK, ankyrin domain; COR, C-terminal of Ras of complex proteins; LRR, leucine-rich repeat domain. (B) Scheme reproducing the lrrk2 gene targeting and screening strategy. The lrrk2 open reading frame (ORF) is highlighted in black; F1, R1: lrrk2 ORF-specific primers; F2, R2: lrrk2 ORF-flanking primers; ts1 (magenta), ts2 (green): gRNA target sites (ts); bp, base pairs. (C) gap-PCR analysis of genomic DNA from wild-type (lrrk2+), heterozygous (lrrk2+/tud113), and homozygous mutant (lrrk2tud113) individuals. F1 and R1 amplify a 289-base pairs-long product, F2 and R2 a 204-base pairs-long product. (D) lrrk2 ISH confirming the complete absence of lrrk2 expression in 22-hpf lrrk2tud113 embryos.
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