CRISPR/Cas9-mediated LRP10 Knockout in HuTu-80 and HEK 293T Cell Lines

Martyna M. Grochowska; Vincenzo Bonifati; Wim Mandemakers

doi:10.21769/BioProtoc.4521

CRISPR/Cas9-mediated LRP10 Knockout in HuTu-80 and HEK 293T Cell Lines

Martyna M. Grochowska^*
, Vincenzo Bonifati
, Wim Mandemakers^*

^*Corresponding author for this work

Clinical Genetics

Research output: Contribution to journal › Article › Academic › peer-review

2 Citations (Scopus)

Abstract

Loss-of-function (LoF) variants in the low-density lipoprotein receptor–related protein 10 gene (LRP10) have been recently implicated in the development of neurodegenerative diseases, including Parkinson's disease (PD), PD dementia (PDD), and dementia with Lewy bodies (DLB). However, despite the genetic evidence, little is known about the LRP10 protein function in health and disease. Here, we describe a detailed protocol to efficiently generate a LRP10 LoF model in two independent LRP10-expressing cell lines, HuTu-80 and HEK 293T, using the CRISPR/Cas9 genome-editing tool. Our method efficiently generates bi-allelic LRP10 knockout (KO), which can be further utilized to elucidate the physiological LRP10 protein function and to model some aspects of neurodegenerative disorders.

Original language	English
Article number	e4521
Journal	Bio-Protocol
Volume	12
Issue number	19
DOIs	https://doi.org/10.21769/BioProtoc.4521
Publication status	Published - 5 Oct 2022

Bibliographical note

Acknowledgments:
This work was supported by research grants from the Stichting Nederland. pSpCas9(BB)-2A-GFP (PX458) was a gift from http://n2t.net/addgene:48138; RRID:Addgene_48138).

Publisher Copyright:
© 2022 The Authors; exclusive licensee Bio-protocol LLC.

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10.21769/BioProtoc.4521

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9548521

Cite this

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title = "CRISPR/Cas9-mediated LRP10 Knockout in HuTu-80 and HEK 293T Cell Lines",

abstract = "Loss-of-function (LoF) variants in the low-density lipoprotein receptor–related protein 10 gene (LRP10) have been recently implicated in the development of neurodegenerative diseases, including Parkinson's disease (PD), PD dementia (PDD), and dementia with Lewy bodies (DLB). However, despite the genetic evidence, little is known about the LRP10 protein function in health and disease. Here, we describe a detailed protocol to efficiently generate a LRP10 LoF model in two independent LRP10-expressing cell lines, HuTu-80 and HEK 293T, using the CRISPR/Cas9 genome-editing tool. Our method efficiently generates bi-allelic LRP10 knockout (KO), which can be further utilized to elucidate the physiological LRP10 protein function and to model some aspects of neurodegenerative disorders.",

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AU - Bonifati, Vincenzo

AU - Mandemakers, Wim

N1 - Acknowledgments: This work was supported by research grants from the Stichting Nederland. pSpCas9(BB)-2A-GFP (PX458) was a gift from http://n2t.net/addgene:48138; RRID:Addgene_48138). Publisher Copyright: © 2022 The Authors; exclusive licensee Bio-protocol LLC.

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N2 - Loss-of-function (LoF) variants in the low-density lipoprotein receptor–related protein 10 gene (LRP10) have been recently implicated in the development of neurodegenerative diseases, including Parkinson's disease (PD), PD dementia (PDD), and dementia with Lewy bodies (DLB). However, despite the genetic evidence, little is known about the LRP10 protein function in health and disease. Here, we describe a detailed protocol to efficiently generate a LRP10 LoF model in two independent LRP10-expressing cell lines, HuTu-80 and HEK 293T, using the CRISPR/Cas9 genome-editing tool. Our method efficiently generates bi-allelic LRP10 knockout (KO), which can be further utilized to elucidate the physiological LRP10 protein function and to model some aspects of neurodegenerative disorders.

AB - Loss-of-function (LoF) variants in the low-density lipoprotein receptor–related protein 10 gene (LRP10) have been recently implicated in the development of neurodegenerative diseases, including Parkinson's disease (PD), PD dementia (PDD), and dementia with Lewy bodies (DLB). However, despite the genetic evidence, little is known about the LRP10 protein function in health and disease. Here, we describe a detailed protocol to efficiently generate a LRP10 LoF model in two independent LRP10-expressing cell lines, HuTu-80 and HEK 293T, using the CRISPR/Cas9 genome-editing tool. Our method efficiently generates bi-allelic LRP10 knockout (KO), which can be further utilized to elucidate the physiological LRP10 protein function and to model some aspects of neurodegenerative disorders.

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CRISPR/Cas9-mediated LRP10 Knockout in HuTu-80 and HEK 293T Cell Lines

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