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带有人类/小鼠人工染色体的人类细胞系面板。

Panel of human cell lines with human/mouse artificial chromosomes.

机构信息

Division of Genome and Cellular Functions, Department of Molecular and Cellular Biology, Faculty of Medicine, School of Life Science, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan.

Chromosome Engineering Research Center, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503, Japan.

出版信息

Sci Rep. 2022 Feb 22;12(1):3009. doi: 10.1038/s41598-022-06814-3.

DOI:10.1038/s41598-022-06814-3
PMID:35194085
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8863800/
Abstract

Human artificial chromosomes (HACs) and mouse artificial chromosomes (MACs) are non-integrating chromosomal gene delivery vectors for molecular biology research. Recently, microcell-mediated chromosome transfer (MMCT) of HACs/MACs has been achieved in various human cells that include human immortalised mesenchymal stem cells (hiMSCs) and human induced pluripotent stem cells (hiPSCs). However, the conventional strategy of gene introduction with HACs/MACs requires laborious and time-consuming stepwise isolation of clones for gene loading into HACs/MACs in donor cell lines (CHO and A9) and then transferring the HAC/MAC into cells via MMCT. To overcome these limitations and accelerate chromosome vector-based functional assays in human cells, we established various human cell lines (HEK293, HT1080, hiMSCs, and hiPSCs) with HACs/MACs that harbour a gene-loading site via MMCT. Model genes, such as tdTomato, TagBFP2, and ELuc, were introduced into these preprepared HAC/MAC-introduced cell lines via the Cre-loxP system or simultaneous insertion of multiple gene-loading vectors. The model genes on the HACs/MACs were stably expressed and the HACs/MACs were stably maintained in the cell lines. Thus, our strategy using this HAC/MAC-containing cell line panel has dramatically simplified and accelerated gene introduction via HACs/MACs.

摘要

人类人工染色体（HACs）和小鼠人工染色体（MACs）是用于分子生物学研究的非整合染色体基因递送载体。最近，HACs/MACs 的微细胞介导染色体转移（MMCT）已在各种人类细胞中实现，包括人类永生化间充质干细胞（hiMSCs）和人类诱导多能干细胞（hiPSCs）。然而，用 HACs/MACs 进行基因导入的传统策略需要在供体细胞系（CHO 和 A9）中进行繁琐且耗时的逐步分离克隆，以将基因加载到 HACs/MACs 中，然后通过 MMCT 将 HAC/MAC 转移到细胞中。为了克服这些限制并加速人类细胞中基于染色体载体的功能测定，我们通过 MMCT 建立了各种带有基因加载位点的人类细胞系（HEK293、HT1080、hiMSCs 和 hiPSCs）。通过 Cre-loxP 系统或同时插入多个基因加载载体，将模型基因（如 tdTomato、TagBFP2 和 ELuc）导入这些预先制备的 HAC/MAC 导入细胞系中。HACs/MACs 上的模型基因稳定表达，并且 HACs/MACs 在细胞系中稳定维持。因此，我们使用这种包含 HAC/MAC 的细胞系面板的策略极大地简化和加速了通过 HACs/MAC 进行的基因导入。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/c6e51c622e98/41598_2022_6814_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/8b704fc00228/41598_2022_6814_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/5749c7bcff58/41598_2022_6814_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/d951a72c2372/41598_2022_6814_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/43542c7de610/41598_2022_6814_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/c6e51c622e98/41598_2022_6814_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/8b704fc00228/41598_2022_6814_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/5749c7bcff58/41598_2022_6814_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/d951a72c2372/41598_2022_6814_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/43542c7de610/41598_2022_6814_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/47f4/8863800/c6e51c622e98/41598_2022_6814_Fig5_HTML.jpg

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