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利用染色质免疫沉淀测序（ChIP-seq）对转基因马-达比牛肾细胞中噬菌体φC31整合酶的结合位点进行全基因组图谱绘制。

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

作者信息

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

机构信息

Department of Laboratory Medicine, Shanghai Eighth People's Hospital, Shanghai, 200040 China.

Department of Respiratory Medicine, Shanghai First People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, 201620 China.

出版信息

Hereditas. 2019 Jan 14;156:3. doi: 10.1186/s41065-018-0079-z. eCollection 2019.

DOI:10.1186/s41065-018-0079-z
PMID:30675136
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6332687/
Abstract

BACKGROUND

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 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.

RESULTS

In 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.

CONCLUSION

This 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.

摘要

背景

ΦC31整合酶是一种位点特异性重组酶，能够有效地将携带attB的转基因靶向到哺乳动物基因组内的内源性假attP位点。内源性结合位点的序列特征将有助于我们全面了解ΦC31整合酶的位点特异性识别功能。本研究旨在揭示牛细胞中ΦC31整合酶结合位点的全局图谱，并通过综合生物信息学方法分析这些结合位点的特征。

结果

在本研究中，我们构建了一种ChIP-seq方法，可用于揭示phiC31整合酶的全局结合位点。共鉴定出6740个潜在的ΦC31整合酶结合位点。发现了一个包含反向重复序列且与野生型attP位点相似的序列基序。使用REPEATMASKER，我们为6740个结合位点共鉴定出20183个分布在50种重复类型中的重复区域。这些位点在生物过程的GO类别“GTPase活性的调节”和信号跨膜转运蛋白活性的KEGG途径中富集。

结论

本研究首次使用ChIP测序方法揭示了ΦC31整合酶结合位点的全局图谱，并分析了这些结合位点的特征。该方法将有助于我们全面了解phiC31整合酶的位点特异性整合功能机制，并可能促进其在基因治疗和转基因动物生成中的基因操作。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/fce50095369f/41065_2018_79_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/8ead81dde93c/41065_2018_79_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/e72fbd32aa47/41065_2018_79_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/cd3891335c7b/41065_2018_79_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/a8cdccc336e9/41065_2018_79_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/747b677ca300/41065_2018_79_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/fce50095369f/41065_2018_79_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/8ead81dde93c/41065_2018_79_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/e72fbd32aa47/41065_2018_79_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/cd3891335c7b/41065_2018_79_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/a8cdccc336e9/41065_2018_79_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/747b677ca300/41065_2018_79_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/e69e/6332687/fce50095369f/41065_2018_79_Fig6_HTML.jpg

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