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Updated: Jul 27, 2026
Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9
Published on: January 3, 2015
Generation of Monosomy 21q Human iPS Cells by CRISPR/Cas9-Mediated Interstitial Megabase Deletion
Masaya Egawa1, Narumi Uno1, Rina Komazaki1
1Laboratory of Bioengineering, School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
Abstract:
Missing an entire chromosome or chromosome arm in normal diploid cells has a deleterious impact on cell viability, which may contribute to the development of specific birth defects. Nevertheless, the effects of chromosome loss in human cells have remained unexplored due to the lack of suitable model systems. Here, we developed an efficient, selection-free approach to generate partial monosomy in human induced pluripotent stem cells (iPSCs). The introduction of Cas9 proteins and a pair of gRNAs induces over megabase-sized interstitial chromosomal deletions. Using human chromosome 21 (HSA21) as a model, partial monosomy 21q (PM21q) iPSC lines with deletions ranging from 4.5 to 27.9 Mb were isolated. A 33.6 Mb deletion, encompassing all protein-coding genes on 21q, was also achieved, establishing the first 21q monosomy human iPSC line. Transcriptome and proteome analyses revealed that the abundances of mRNA and protein encoded by the majority of genes in the monosomic regions are half of the diploid expression level, indicating an absence of dosage compensation. The ability to generate customized partial monosomy cell lines on an isogenic, karyotypically normal background should facilitate the gain of novel insights into the impact of chromosome loss on cellular fitness.
Insights
Researchers created a new method to generate partial monosomy in human stem cells, enabling the study of chromosome loss effects. This breakthrough provides the first human iPSC line with complete 21q monosomy, revealing no dosage compensation.
Area of Science:
- Genetics
- Stem Cell Biology
- Genomics
Background:
- Chromosome loss in diploid cells can impact cell viability and contribute to birth defects.
- Studying chromosome loss effects in human cells is challenging due to a lack of suitable model systems.
Purpose of the Study:
- To develop an efficient, selection-free method for generating partial monosomy in human induced pluripotent stem cells (iPSCs).
- To create the first human iPSC line exhibiting complete 21q monosomy.
- To investigate the impact of chromosome loss on gene expression and cellular fitness.
Main Methods:
- Utilized Cas9 proteins and guide RNAs (gRNAs) to induce large interstitial chromosomal deletions in human iPSCs.
- Generated partial monosomy 21q (PM21q) iPSC lines with deletions ranging from 4.5 to 33.6 Mb.
- Performed transcriptome and proteome analyses to assess gene expression levels.
Main Results:
- Successfully generated iPSC lines with partial and complete deletions on chromosome 21q.
- Demonstrated that genes within monosomic regions exhibit approximately half the diploid expression level.
- Confirmed the absence of dosage compensation in the generated monosomic cell lines.
Conclusions:
- The developed method allows for the creation of customized partial monosomy cell lines on an isogenic background.
- These iPSC models facilitate novel insights into the cellular consequences of chromosome loss.
- The findings highlight the lack of dosage compensation in human cells with partial or complete chromosome arm loss.
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