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In vivo CRISPR screening protocol to identify metastasis mediators using iteratively selected mouse models.

Yuqi Wang1, Mangze Hu2, Yilong Zou1

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This study details an in vivo CRISPR screening protocol to identify genes driving ovarian cancer metastasis in living organisms. This method overcomes limitations of lab-based models for discovering critical metastasis genes.

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Area of Science:

  • Genetics
  • Oncology
  • Molecular Biology

Background:

  • In vitro models have limitations in fully recapitulating complex biological processes like cancer metastasis.
  • Identifying genes that drive metastasis in native physiological contexts is crucial for understanding and treating advanced cancers.

Purpose of the Study:

  • To present a detailed protocol for an in vivo CRISPR screening technique to identify metastasis-driving genes in ovarian cancer.
  • To provide a reproducible method for researchers studying cancer progression and metastasis.

Main Methods:

  • The protocol involves single-guide RNA (sgRNA) library design and validation.
  • Establishment of metastatic mouse models and subsequent tissue collection are key steps.
  • Bioinformatics analysis of sgRNA amplification by sequencing is used for candidate gene identification.

Main Results:

  • The in vivo CRISPR screen successfully identifies potential metastasis-driving genes within the native tumor microenvironment.
  • The protocol enables the discovery of genes that may be missed by traditional in vitro screening methods.

Conclusions:

  • This in vivo CRISPR screening protocol offers a powerful approach to uncover novel metastasis genes in ovarian cancer.
  • The methodology surpasses in vitro limitations, providing insights into metastasis in a more relevant biological context.