ArticleNucleic acids research2010
BAC TG-EMBED: one-step method for high-level, copy-number-dependent, position-independent transgene expression.
Article in Nucleic acids research, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed.
- Acidic transcription factors position the genome at nuclear speckles through transcription-dependent and -independent mechanisms.Molecular cell · 2026Article
- BAC Recombineering and Transgenesis to Study Cell Polarity and Polarized Tissue Morphogenesis in Mice.Methods in molecular biology (Clifton, N.J.) · 2022Article
- Efficient and Reproducible Multigene Expression after Single-Step Transfection Using Improved BAC Transgenesis and Engineering Toolkit.ACS synthetic biology · 2020Article
- Stable and reproducible transgene expression independent of proliferative or differentiated state using BAC TG-EMBED.Gene therapy · 2018Article
- Altered Baseline and Nicotine-Mediated Behavioral and Cholinergic Profiles in ChAT-Cre Mouse Lines.The Journal of neuroscience : the official journal of the Society for Neuroscience · 2018Article
- Comparative Analysis of piggyBac, CRISPR/Cas9 and TALEN Mediated BAC Transgenesis in the Zygote for the Generation of Humanized SIRPA Rats.Scientific reports · 2016Article
- Single cell visualization of transcription kinetics variance of highly mobile identical genes using 3D nanoimaging.Scientific reports · 2015Article
- Human histone acetyltransferase 1 (Hat1) acetylates lysine 5 of histone H2A in vivo.Molecular and cellular biochemistry · 2014Article
- Recombineering strategies for developing next generation BAC transgenic tools for optogenetics and beyond.Frontiers in behavioral neuroscience · 2014Article
- β-Globin cis-elements determine differential nuclear targeting through epigenetic modifications.The Journal of cell biology · 2013Article
- Transgenically mediated shRNAs targeting conserved regions of foot-and-mouth disease virus provide heritable resistance in porcine cell lines and suckling mice.Veterinary research · 2013Article
- Reliable transgene-independent method for determining Sleeping Beauty transposon copy numbers.Mobile DNA · 2011Article
- Recent advances in recombinant protein production: BAC-based expression vectors, the bigger the better.BioengineeredArticle
Corrections and comments
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Authors and funding
2 authors.
Funding
Abstract
Chromosome position effects combined with transgene silencing of multi-copy plasmid insertions lead to highly variable and usually quite low expression levels of mini-genes integrated into mammalian chromosomes. Together, these effects greatly complicate obtaining high-level expression of therapeutic proteins in mammalian cells or reproducible expression of individual or multiple transgenes. Here, we report a simple, one-step procedure for obtaining high-level, reproducible mini-gene expression in mammalian cells. By inserting mini-genes at different locations within a BAC containing the DHFR housekeeping gene locus, we obtain copy-number-dependent, position-independent expression with chromosomal insertions of one to several hundred BAC copies. These multi-copy DHFR BAC insertions adopt similar large-scale chromatin conformations independent of their chromosome integration site, including insertions within centromeric heterochromatin. Prevention of chromosome position effects, therefore, may be the result of embedding the mini-gene within the BAC-specific large-scale chromatin structure. The expression of reporter mini-genes can be stably maintained during continuous, long-term culture in the presence of drug selection. Finally, we show that this method is extendable to reproducible, high-level expression of multiple mini-genes, providing improved expression of both single and multiple transgenes.
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