Gene Functional Studies Using Bacterial Artificial Chromosome (BACs)

A Scalable Epitope Tagging Approach for High Throughput ChIP-Seq Analysis

“…9 However, genetic engineering of BACs can be time-consuming and laborious. 10 Alternatively, adeno-associated virus (rAAV) has been employed as a delivery vehicle for a knock-in (KI) vector to realize the epitope tag insertion and overcome low recombination efficiency. 11,12 More recently, a method was reported that directly modifies an endogenous locus via CRISPR/Cas9 mediated homologous recombination (HR).…”

“…Genome-wide profiling of TF-DNA interactions is crucial for dissecting the transcriptional regulation networks that govern the spatiotemporal expression of genes in an organism. − Chromatin immunoprecipitation followed by sequencing (ChIP-Seq) is the most common method for this purpose. − However, this technique is limited by the availability of ChIP-grade antibodies against transcription factors. , To address this bottleneck, bacterial artificial chromosome (BAC) vector has been employed to introduce epitope tags to transcription factors for ChIP-Seq experiments by highly specific antibodies against the epitopes . However, genetic engineering of BACs can be time-consuming and laborious . Alternatively, adeno-associated virus (rAAV) has been employed as a delivery vehicle for a knock-in (KI) vector to realize the epitope tag insertion and overcome low recombination efficiency. , More recently, a method was reported that directly modifies an endogenous locus via CRISPR/Cas9 mediated homologous recombination (HR) .…”

“…10.1021/acssynbio.6b00358. Supplementary methods, additional figures and tables (PDF) Phone: 1-(858)-822-5766.…”