One-step generation of triple gene-targeted pigs using CRISPR/Cas9 system

Wang, Xianlong; Cao, Chunwei; Huang, Jiaojiao; Yao, Jing; Hai, Tang; Zheng, Qiantao; Wang, Xiao; Zhang, Hongyong; Qin, Guosong; Cheng, Jinbo; Wang, Yanfang; Yuan, Zengqiang; Zhou, Qi; Wang, Hongmei; Zhao, Jianguo

doi:10.1038/srep20620

Cited by 111 publications

(81 citation statements)

References 24 publications

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“…In addition, pigs, especially minipigs, also share a number of similarities with human, such as body size, anatomy, physiology, pathology as well as genome, providing an excellent animal model for mimicking human diseases (Nagashima et al, ; Niemann & Lucas‐Hahn, ; Prather et al, ; Whyte & Prather, ). Genetically modified pigs have also been widely used for modelling numerous human diseases, such as Alzheimer’s disease (Kragh et al, ; Lee et al, ), Huntington’s disease (Yan et al, ; Yang et al, ) and Parkinson’s disease (Wang et al, ; Zhou et al, ). Therefore, genetically modified pigs hold great promise in the fields of agriculture and biomedicine.…”

Section: Discussionmentioning

confidence: 99%

Generation of transgenic‐cloned Huanjiang Xiang pigs systemically expressing enhanced green fluorescent protein

Zhu

Zhong²,

Ge³

et al. 2018

Reprod Domestic Animals

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Huanjiang Xiang pig is a unique native minipig breed originating in Guangxi, China, and has great utility value in agriculture and biomedicine. Reproductive biotechnologies such as somatic cell nuclear transfer (SCNT) and SCNT-mediated genetic modification show great potential value in genetic preservation and utilization of Huanjiang Xiang pigs. Our previous work has successfully produced cloned and transgenic-cloned embryos using somatic cells from a Huanjiang Xiang pig. In this study, we firstly report the generation of transgenic-cloned Huanjiang Xiang pigs carrying an enhanced green fluorescent protein (eGFP) gene. A total of 504 SCNT-derived embryos were transferred to two surrogate recipients, one of which became pregnant and gave birth to three live piglets. Exogenous eGFP transgene had integrated in all of the three Huanjiang Xiang piglets identified by genotyping. Furthermore, expression of eGFP was also detected from in vitro cultured skin fibroblast cells and various organs or tissues from positive transgenic-cloned Huanjiang Xiang pigs. The present work provides a practical method to preserve this unique genetic resource and also lays a foundation for genetic modification of Huanjiang Xiang pigs with improved values in agriculture and biomedicine.

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Section: Discussionmentioning

confidence: 99%

Generation of transgenic‐cloned Huanjiang Xiang pigs systemically expressing enhanced green fluorescent protein

Zhu

Zhong²,

Ge³

et al. 2018

Reprod Domestic Animals

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“…These delivery systems can be classified as non‐virus and virus. Non‐virus delivery systems include electroporation or nucleofection, lipid‐based transfection, cationic peptides, and microinjection (Cockrell & Kafri, ; Komor et al, ; Luo & Saltzman, ; Maasho, Marusina, Reynolds, Coligan, & Borrego, ; Moreno & Mali, ; Wang et al, ; Yin, Kanasty, et al, ; Zeitelhofer et al, ). Virus delivery systems rely on natural viral vectors, such as retroviruses, lentiviruses, adenoviruses, and adeno‐associated viruses (AAVs).…”

Section: Improved Understanding Of Crispr/cas Systemsmentioning

confidence: 99%

CRISPR editing in biological and biomedical investigation

Huang

Wang

Zhao

2017

Journal Cellular Physiology

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Recently, clustered regularly interspaced short palindromic repeats (CRISPR) based genomic editing technologies have armed researchers with powerful new tools to biological and biomedical investigations. To further improve and expand its functionality, natural, and engineered CRISPR associated nine proteins (Cas9s) have been investigated, various CRISPR delivery strategies have been tested and optimized, and multiple schemes have been developed to ensure precise mammalian genome editing. Benefiting from those in-depth understanding and further development of CRISPR, versatile CRISPR-based platforms for genome editing have been rapidly developed to advance investigations in biology and biomedicine. In biological research area, CRISPR has been widely adopted in both fundamental and applied research fields, such as accurate base editing, transcriptional regulation, and genome-wide screening. In biomedical research area, CRISPR has also shown its extensive applicability in the establishment of animal models for genetic disorders especially those large animals and non-human primates models, and gene therapy to combat virus infectious diseases, to correct monogenic disorders in vivo or in pluripotent cells. In this prospect article, after highlighting recent developments of CRISPR systems, we outline different applications and current limitations of CRISPR use in biological and biomedical investigation. Finally, we provide a perspective for future development and potential risks of this multifunctional technology.

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“…Later, TALENs were used to knockout the alleles of LDL receptor (LDLR) gene in swine. Recently, CRISPR/Cas9 system was applied for genome editing in pigs (Hai et al, 2014;Whitworth et al, 2014;Wang et al, 2016).…”

Section: Livestockmentioning

confidence: 99%

Programmable Site-Specific Nucleases for Targeted Genome Engineering in Higher Eukaryotes

Govindan

Ramalingam

2016

J. Cell. Physiol.

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Recent advances in the targeted genome engineering enable molecular biologists to generate sequence specific modifications with greater efficiency and higher specificity in complex eukaryotic genomes. Programmable site-specific DNA cleavage reagents and cellular DNA repair mechanisms have made this possible. These reagents have become powerful tools for delivering a site-specific genomic double-strand break (DSB) at the desired chromosomal locus, which produces sequence alterations through error-prone non-homologous end joining (NHEJ) resulting in gene inactivations/knockouts. Alternatively, the DSB can be repaired through homology-directed repair (HDR) using a donor DNA template, which leads to the introduction of desired sequence modifications at the predetermined site. Here, we summarize the role of three classes of nucleases; zinc finger nucleases (ZFNs), transcription activator like effector nucleases (TALENs), and clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) system in achieving targeted genome modifications. Further, we discuss the progress towards the applications of programmable site-specific nucleases (SSNs) in treating human diseases and other biological applications in economically important higher eukaryotic organisms such as plants and livestock. J. Cell. Physiol. 231: 2380-2392, 2016. © 2016 Wiley Periodicals, Inc.

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One-step generation of triple gene-targeted pigs using CRISPR/Cas9 system

Cited by 111 publications

References 24 publications

Generation of transgenic‐cloned Huanjiang Xiang pigs systemically expressing enhanced green fluorescent protein

Generation of transgenic‐cloned Huanjiang Xiang pigs systemically expressing enhanced green fluorescent protein

CRISPR editing in biological and biomedical investigation

Programmable Site-Specific Nucleases for Targeted Genome Engineering in Higher Eukaryotes

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