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In-vitro Mutagenesis

In-vitro Mutagenesis

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.

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Updated: Jun 21, 2025

Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells

Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells

Published on: June 16, 2017

Optimized CRISPR-based knockout in BeWo cells.

Eric Yin¹, Meagan N Esbin¹

¹University of California, Berkeley, United States.

|July 13, 2024

Summary

This summary is machine-generated.

This study presents an optimized CRISPR-Cas9 protocol for efficient gene knockout in BeWo cells, a model for placental research. The method uses ribonucleoprotein complexes and electroporation, improving genetic manipulation in these challenging cells.

Keywords:

CRISPR Cas9 Cell culture Cell lines DNA expression and control Fusion/differentiation Knockout Syncytiotrophoblast Transfection

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Last Updated: Jun 21, 2025

Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells

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Published on: June 16, 2017

Highly Efficient Gene Disruption of Murine and Human Hematopoietic Progenitor Cells by CRISPR/Cas9

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Electroporation-Based CRISPR-Cas9-Mediated Gene Knockout in THP-1 Cells and Single-Cell Clone Isolation

Electroporation-Based CRISPR-Cas9-Mediated Gene Knockout in THP-1 Cells and Single-Cell Clone Isolation

Published on: February 28, 2025

Area of Science:

Molecular Biology
Genetics
Cell Biology

Background:

CRISPR genome editing is crucial for genotype-phenotype studies.
Efficient editing is challenging in certain cell types like BeWo cells, hindering research on placental syncytiotrophoblast function.

Purpose of the Study:

To develop and optimize an easy-to-use CRISPR-Cas9 protocol for gene knockout in BeWo cells.
To provide guidelines for guide RNA design and knockout assessment in this cell line.

Main Methods:

Utilized CRISPR-Cas9 ribonucleoprotein (RNP) complexes delivered via electroporation for gene editing.
Established parameters for guide RNA design and validated the protocol using Syncytin-2 (ERVFRD-1) knockout.

Main Results:

Achieved highly efficient gene knockout in BeWo cells using the optimized RNP electroporation protocol.
Demonstrated successful assessment of genetic editing efficiency at the Syncytin-2 locus.

Conclusions:

The developed protocol facilitates efficient RNP-mediated genetic knockouts in BeWo cells, aiding the study of placental cell-cell fusion and hormone secretion.
This optimized nucleofection and RNP delivery strategy may benefit editing in other difficult trophoblast cells or for transgene delivery.