Panel of human cell lines with human/mouse artificial chromosomes

Area of Science:

• Molecular Biology
• Genetics
• Cell Biology

Background:

• Human artificial chromosomes (HACs) and mouse artificial chromosomes (MACs) are non-integrating gene delivery vectors.
• Microcell-mediated chromosome transfer (MMCT) enables HAC/MAC transfer into human cells, including stem cells.
• Conventional gene introduction using HACs/MACs is laborious and time-consuming.

Purpose of the Study:

• To overcome limitations of conventional HAC/MAC gene introduction methods.
• To accelerate chromosome vector-based functional assays in human cells.
• To establish human cell lines pre-equipped with HACs/MACs harboring a gene-loading site.

Main Methods:

• Established human cell lines (HEK293, HT1080, hiMSCs, hiPSCs) containing HACs/MACs with a gene-loading site via MMCT.
• Introduced model genes (tdTomato, TagBFP2, ELuc) into these cell lines using the Cre-loxP system or simultaneous gene-loading vectors.
• Assessed stable gene expression and chromosome maintenance.

Main Results:

• Successfully created various human cell lines harboring HACs/MACs with a gene-loading site.
• Demonstrated stable expression of model genes introduced into the HACs/MACs.
• Confirmed stable maintenance of HACs/MACs within the recipient human cell lines.

Conclusions:

• The developed strategy significantly simplifies and accelerates gene introduction via HACs/MACs.
• The panel of HAC/MAC-containing cell lines provides a powerful tool for molecular biology research.
• This approach facilitates rapid functional assays using chromosome vectors in human cells.