title: Targeted editing of goat genome with modular-assembly zinc finger nucleases based on activity prediction by computational molecular modeling.
abstract: Zinc finger nuclease (ZFN) technology can mediate targeted genome modification to produce transgenic animals in a high-efficient and biological-safe way. Modular assembly is a rapid, convenient and open-source method for the synthesis of ZFNs. However, this biotechnology is hampered by multistep construction, low-efficiency editing and off-target cleavage. Here we synthesized and tested six pairs of three- or four-finger ZFNs to target one site in goat beta-lactoglobulin (BLG, a dominant allergen in goat milk) gene. Homology modeling was applied to build the structure model of ZFNs to predict their editing activities targeting at goat BLG gene. Goat fibroblast cells were transfected with plasmids that encoded ZFN pairs, and genomic DNA was isolated 72 h later for genome editing efficiency assay. The results of editing efficiency assay demonstrated that ZFNs with optimal interaction modes can edit goat BLG gene more efficiently, whereas ZFNs with unexpected interaction modes showed lower activities in editing BLG gene. We concluded that modular-assembly ZFNs can provide a rapid, public-available, and easy-to-practice platform for transgenic animal research and molecular modeling would help as a useful tool for ZFNs activity prediction.
authors: College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, 210095, People's Republic of China.
authors: Kai
authors: K
authors: Xiong
authors: Shanshan
authors: S
authors: Li
authors: Hongxiao
authors: H
authors: Zhang
authors: Ye
authors: Y
authors: Cui
authors: Debing
authors: D
authors: Yu
authors: Yan
authors: Y
authors: Li
authors: Wenxing
authors: W
authors: Sun
authors: Yingying
authors: Y
authors: Fu
authors: Yun
authors: Y
authors: Teng
authors: Zhi
authors: Z
authors: Liu
authors: Xiaolong
authors: X
authors: Zhou
authors: Peng
authors: P
authors: Xiao
authors: Juan
authors: J
authors: Li
authors: Honglin
authors: H
authors: Liu
authors: Jie
authors: J
authors: Chen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019295:Computational Biology / Q000379:methods; D004720:Endonucleases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D016678:Genome; D023281:Genomics; D006041:Goats; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016415:Sequence Alignment; D016335:Zinc Fingers / Q000235:genetics*
chemical_list: D004720:Endonucleases
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