CRISPR/Cas9 Knockout of Shell Matrix Protein 1 in the Slipper-Snail Crepidula atrasolea

Area of Science:

• Biomineralization research
• Marine biology
• Developmental biology

Background:

• Molluscan shells exhibit remarkable diversity in structure and patterning.
• Shell matrix proteins (SMPs) are crucial for biomineralization, influencing shell formation and morphology.
• Directly studying SMP function in molluscs was previously limited by a lack of gene-editing tools.

Purpose of the Study:

• To report the first successful application of CRISPR/Cas9 gene editing to target a shell matrix protein (SMP) in gastropod molluscs.
• To establish Crepidula atrasolea as a model organism for genetic studies of shell formation.
• To provide a methodological framework for future CRISPR/Cas9 applications in other molluscan species.

Main Methods:

• CRISPR/Cas9 gene editing was employed to generate knockouts of the SMP1 gene in Crepidula atrasolea.
• Gene editing success was verified using Sanger and MiSeq sequencing.
• Loss of SMP1 expression was confirmed through high-content imaging of crispant embryos.

Main Results:

• The study successfully generated SMP1 gene knockouts in the gastropod Crepidula atrasolea.
• CRISPR/Cas9 mediated gene editing was confirmed through molecular sequencing and embryo imaging.
• This demonstrates the efficacy of gene editing for studying SMP function in molluscs.

Conclusions:

• CRISPR/Cas9 technology is effectively applicable to gastropod molluscs for studying biomineralization.
• Crepidula atrasolea is established as a valuable model for investigating the genetic underpinnings of shell formation.
• This research enables future studies to elucidate the precise roles of SMPs in shaping diverse molluscan shell structures.