Improving the Efficiency of Gene Insertion in a Human Artificial Chromosome Vector and Its Transfer in Human-induced Pluripotent Stem Cells

Overview

Journal Biol Methods Protoc

Publisher Oxford University Press

Specialty Biology

Date 2020 Mar 13

PMID 32161806

Citations 7

Authors

Yoshinori Hasegawa

Masashi Ikeno

Nobutaka Suzuki

Manabu Nakayama

Osamu Ohara

Affiliations

Soon will be listed here.

Abstract

A human artificial chromosome (HAC) vector has potential to overcome the problems of stable gene expression associated with plasmid, transposon, and virus-based vectors, such as insertional mutagenesis, position effect, uncontrollable copy number, unstable gene expression, and DNA size limitation. The main advantages of the HAC are its episomal nature and ability to accommodate DNA inserts of any size. However, HAC vectors have two disadvantages: low efficiency of gene insertion and lack of reports regarding the successful HAC transfer to human-induced pluripotent stem cells (iPSCs). We here provide the first report of a method for the efficient transfer of HAC to human iPSCs for obtaining reproducible experimental results. Moreover, we achieved a 10% increase in the gene insertion efficiency in the HAC vector using our new site-specific recombination systems VCre/VloxP and SCre/SloxP.

Citing Articles

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Treatment of CHO cells with Taxol and reversine improves micronucleation and microcell-mediated chromosome transfer efficiency.

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