Global mapping of binding sites for phic31 integrase in transgenic maden-darby bovine kidney cells using ChIP-seq

Lijuan Qu,Guoting Li,Aying Ma,Shuwu Xie,Yue Dai,Fengying Sheng,Qiang Ou,Xueyuan Zhu,Yan Zhang,Xiaoqing Wei,Lei Wang

doi:10.1186/s41065-018-0079-z

Abstract

BackgroundΦC31 integrase, a site-specific recombinase, can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes. The sequence features of endogenous binding sites will help us to fully understand the site-specific recognition function by ΦC31 integrase. The present study was aimed to uncover the global map of ΦC31 integrase binding sites in bovine cells and analysis the features of these binding sites by comprehensive bioinformatics methods.ResultsIn this study, we constructed a ChIP-seq method that can be used to uncover the global binding sites by phiC31 integrase. 6740 potential ΦC31 integrase binding sites were identified. A sequence motif was found that contains inverted repeats and has similarities to wild-type attP site. Using REPEATMASKER, we identified a total of 20,183 repeat-regions distributed in 50 repeat types for the 6740 binding sites. These sites enriched in “regulation of GTPase activity” of in the GO category of biological process and KEGG pathway of signal transmembrane transporter activity.ConclusionThis study is the first time to uncover the global map of binding sites for ΦC31 integrase using ChIP-sequencing method and analysis the features of these binding sites. This method will help us to fully understand the mechanism of the site-specific integration function by phiC31 integrase and will potentially boost its genetic manipulations in both gene therapy and generation of transgenic animals.

Highlights

ΦC31 integrase, a site-specific recombinase, can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes
AttP and attB are unlikely to occur in the genomes of mammals and most plants, but pseudo attP sites are present in the eukaryotic genome [2,3,4,5], ΦC31 integrase can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes [6, 7]
The BpsM1 site we found in bovine maden-darby bovine kidney (MDBK)cell line was not detected in bovine ear fibroblasts [18]

Summary

Introduction

ΦC31 integrase, a site-specific recombinase, can efficiently target attB-bearing transgenes to endogenous pseudo attP sites within mammalian genomes. The sequence features of endogenous binding sites will help us to fully understand the site-specific recognition function by ΦC31 integrase. The 19q13.31 site in the human genome is detectable in all cell types, and 3q26.31 appears only in HepG2 cells [21] These phenomena indicate that, in addition to sequence features, the recognition and binding of the integrase to the sites are affected by other factors such as chromatin structure. This is still not fully studied and understood. These methods have limitations and provide only a small amount of binding sites information

Methods

Results

Discussion

Conclusion