Generation of gene-modified cynomolgus monkey via Cas9/RNA-mediated gene targeting in one-cell embryos.

Published on Feb 1, 2014in Cell 31.40
· DOI :10.1016/j.cell.2014.01.027
Yuyu Niu15
Estimated H-index: 15
,
Bin Shen14
Estimated H-index: 14
(National Resource Center)
+ 25 AuthorsJiahao Sha32
Estimated H-index: 32
(Nanjing Medical University)
Abstract
Summary Monkeys serve as important model species for studying human diseases and developing therapeutic strategies, yet the application of monkeys in biomedical researches has been significantly hindered by the difficulties in producing animals genetically modified at the desired target sites. Here, we first applied the CRISPR/Cas9 system, a versatile tool for editing the genes of different organisms, to target monkey genomes. By coinjection of Cas9 mRNA and sgRNAs into one-cell-stage embryos, we successfully achieve precise gene targeting in cynomolgus monkeys. We also show that this system enables simultaneous disruption of two target genes ( Ppar-γ and Rag1 ) in one step, and no off-target mutagenesis was detected by comprehensive analysis. Thus, coinjection of one-cell-stage embryos with Cas9 mRNA and sgRNAs is an efficient and reliable approach for gene-modified cynomolgus monkey generation.
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References25
Published on Feb 1, 2012in American Journal of Primatology 2.29
Yongchang Chen12
Estimated H-index: 12
,
Yuyu Niu15
Estimated H-index: 15
+ 9 AuthorsYongqing Lu4
Estimated H-index: 4
(Kunming Institute of Zoology)
Much effort has been focused on improving assisted reproductive technology procedures in humans and nonhuman primates (NHPs). However, the pregnancy rate after embryo transfer (ET) has not been satisfactory, indicating that some barriers still need to be overcome in this important procedure. One of the key factors is embryo–uterine synchronicity, which is little known in NHPs. The objective of this study was to investigate the available ET time window in rhesus monkey (Macaca mulatta). Eighty-tw...
9 Citations Source Cite
Jiankui Zhou5
Estimated H-index: 5
(National Resource Center),
Bin Shen14
Estimated H-index: 14
(National Resource Center)
+ 9 AuthorsBian Hu6
Estimated H-index: 6
(National Resource Center)
Taking advantage of the multiplexable genome engineering feature of the CRISPR/Cas9 system, we sought to generate different kinds of immunodeficient mouse strains by embryo co-microinjection of Cas9 mRNA and multiple sgRNAs targeting mouse B2m, Il2rg, Prf1, Prkdc, and Rag1. We successfully achieved multiple gene modifications, fragment deletion, double knockout of genes localizing on the same chromosome, and got different kinds of immunodeficient mouse models with different heritable genetic mod...
71 Citations Source Cite
Qiang Sun6
Estimated H-index: 6
(East China Normal University),
Juan Dong4
Estimated H-index: 4
(Georgia Regents University)
+ 6 AuthorsJoe Z. Tsien44
Estimated H-index: 44
One of the technical bottlenecks in producing nonhuman primate models is that current assisted reproductive techniques, such as in vitro culture and frozen conservation of multicell-stage embryos, often result in poor embryo quality and subsequently lead to low birth rates. We investigated whether pronuclear embryo transfer can be used as an effective means for improving pregnancy and live birth rates of nonhuman primates. We collected 174 metaphase II oocytes by laparoscopy from 22 superovulate...
17 Citations Source Cite
Published on Nov 6, 2013in Nature 41.58
Helen Shen7
Estimated H-index: 7
14 Citations Source Cite
Yuyu Niu15
Estimated H-index: 15
,
Ying Yu20
Estimated H-index: 20
+ 19 AuthorsWei Si10
Estimated H-index: 10
The development of transgenic technologies in monkeys is important for creating valuable animal models of human physiology so that the etiology of diseases can be studied and potential therapies for their amelioration may be developed. However, the efficiency of producing transgenic primate animals is presently very low, and there are few reports of success. We have developed an improved methodology for the production of transgenic rhesus monkeys, making use of a simian immunodeficiency virus (S...
45 Citations Source Cite
Published on Aug 1, 2011in Nature Biotechnology 35.72
Laurent Tesson29
Estimated H-index: 29
(French Institute of Health and Medical Research),
Claire Usal27
Estimated H-index: 27
(French Institute of Health and Medical Research)
+ 10 AuthorsLei Zhang27
Estimated H-index: 27
(Sangamo BioSciences)
478 Citations Source Cite
Published on Feb 15, 2013in Science 41.06
Prashant Mali28
Estimated H-index: 28
(Harvard University),
Luhan Yang16
Estimated H-index: 16
(Harvard University)
+ 5 AuthorsGeorge M Church G M132
Estimated H-index: 132
(Wyss Institute for Biologically Inspired Engineering)
Bacteria and archaea have evolved adaptive immune defenses, termed clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems, that use short RNA to direct degradation of foreign nucleic acids. Here, we engineer the type II bacterial CRISPR system to function with custom guide RNA (gRNA) in human cells. For the endogenous AAVS1 locus, we obtained targeting rates of 10 to 25% in 293T cells, 13 to 8% in K562 cells, and 2 to 4% in induced pluripotent stem cel...
4,240 Citations Source Cite
Published on Jan 12, 2001in Science 41.06
Anthony W.S. Chan29
Estimated H-index: 29
,
Kowit-Yu Chong17
Estimated H-index: 17
+ 2 AuthorsGerald Schatten70
Estimated H-index: 70
Transgenic rhesus monkeys carrying the green fluorescent protein (GFP) gene were produced by injecting pseudotyped replication-defective retroviral vector into the perivitelline space of 224 mature rhesus oocytes, later fertilized by intracytoplasmic sperm injection. Of the three males born from 20 embryo transfers, one was transgenic when accessible tissues were assayed for transgene DNA and messenger RNA. All tissues that were studied from a fraternal set of twins, miscarried at 73 days, carri...
236 Citations Source Cite
Published on Aug 1, 2013in Nature Biotechnology 35.72
Wei Li10
Estimated H-index: 10
,
Fei Teng7
Estimated H-index: 7
+ 1 AuthorsQi Zhou41
Estimated H-index: 41
Simultaneous generation and germline transmission of multiple gene mutations in rat using CRISPR-Cas systems
276 Citations Source Cite
Published on Jun 1, 2008in Nature 41.58
Shang Hsun Yang16
Estimated H-index: 16
,
Pei Hsun Cheng9
Estimated H-index: 9
(Emory University)
+ 14 AuthorsJack Orkin1
Estimated H-index: 1
(Emory University)
Huntington's disease is a severely disabling and lethal neurodegenerative disorder. The development of a non-human primate model for the disease would be invaluable in understanding its pathology and in developing therapeutic strategies, and this paper reports a significant step towards that goal is reported in this issue. Transgenic rhesus macaque monkeys that express the first exon of the polyglutamine-expanded human huntingtin gene develop key features of Huntington's disease, including dysto...
325 Citations Source Cite
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Cited By566
Published on Jan 1, 2014in Methods in Enzymology 1.98
Benjamin L. Oakes8
Estimated H-index: 8
(University of California, Berkeley),
Dana C. Nadler5
Estimated H-index: 5
(University of California, Berkeley),
David F. Savage19
Estimated H-index: 19
(University of California, Berkeley)
Abstract CRISPR/Cas systems act to protect the cell from invading nucleic acids in many bacteria and archaea. The bacterial immune protein Cas9 is a component of one of these CRISPR/Cas systems and has recently been adapted as a tool for genome editing. Cas9 is easily targeted to bind and cleave a DNA sequence via a complementary RNA; this straightforward programmability has gained Cas9 rapid acceptance in the field of genetic engineering. While this technology has developed quickly, a number of...
8 Citations Source Cite
Published on Sep 1, 2015in Hearing Research 2.82
Bing Zou4
Estimated H-index: 4
(University of Miami),
Rahul Mittal11
Estimated H-index: 11
(University of Miami)
+ 9 AuthorsShiming Yang13
Estimated H-index: 13
(Chinese PLA General Hospital)
Abstract Targeted genome editing mediated by clustered, regularly interspaced, short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9 (Cas9) technology has emerged as one of the most powerful tools to study gene functions, and with potential to treat genetic disorders. Hearing loss is one of the most common sensory disorders, affecting approximately 1 in 500 newborns with no treatment. Mutations of inner ear genes contribute to the largest portion of genetic deafness. The simplicity and ...
17 Citations Source Cite
Published on Jul 1, 2015in Journal of Community Genetics
José Roberto Goldim11
Estimated H-index: 11
(Pontifícia Universidade Católica do Rio Grande do Sul)
Genetics and ethics have had numerous convergences and divergences over time. From Darwin through the Asilomar Conference and the Universal Declaration on the Human Genome and Human Rights, much has been achieved, but much still remains to be done. The use of biological materials that are already being stored and the adequacy of new technologies, such as clustered regularly interspaced short palindromic repeats (CRISPR), are some of the current challenges of this possible and necessary interacti...
1 Citations Source Cite
Published on Oct 1, 2015in Neurobiology of Learning and Memory 3.24
Sandrine Camus7
Estimated H-index: 7
,
Wai Kin D. Ko10
Estimated H-index: 10
+ 1 AuthorsErwan Bezard65
Estimated H-index: 65
(Centre national de la recherche scientifique)
Abstract Although everyone would agree that successful translation of therapeutic candidates for central nervous disorders should involve non-human primate (nhp) models of cognitive disorders, we are left with the paucity of publications reporting either the target validation or the actual preclinical testing in heuristic nhp models. In this review, we discuss the importance of nhps in translational research, highlighting the advances in technological/methodological approaches for ‘bridging the ...
16 Citations Source Cite
Published on Aug 1, 2015in Journal of Genetics and Genomics 4.07
Fengjiao Chen1
Estimated H-index: 1
(Nanjing Medical University),
Ying Wang2
Estimated H-index: 2
(Nanjing Medical University)
+ 16 AuthorsManling Zhang5
Estimated H-index: 5
(Nanjing Medical University)
Generating B cell-deficient mutant is the first step to produce human antibody repertoires in large animal models. In this study, we applied the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) system to target the JH region of the pig IgM heavy chain gene which is crucial for B cell development and differentiation. Transfection of IgM-targeting Cas9 plasmid in primary porcine fetal fibroblasts (PFFs) enabled inducing gene knock out (KO) in up to 53.3% of...
18 Citations Source Cite
Published on Dec 1, 2015in Scientific Reports 4.12
Jingwei Wei2
Estimated H-index: 2
,
Stefan Wagner4
Estimated H-index: 4
+ 4 AuthorsGötz Laible17
Estimated H-index: 17
The recent development of designer nucleases allows for the efficient and precise introduction of genetic change into livestock genomes. Most studies so far have focused on the introduction of random mutations in cultured cells and the use of nuclear transfer to generate animals with edited genotypes. To circumvent the intrinsic uncertainties of random mutations and the inefficiencies of nuclear transfer we directed our efforts to the introduction of specific genetic changes by homology-driven r...
19 Citations Source Cite
Published on Jun 1, 2015in Journal of Genetics and Genomics 4.07
Yong-Gang Yao41
Estimated H-index: 41
(Kunming Institute of Zoology),
Yongbin Chen17
Estimated H-index: 17
(Kunming Institute of Zoology),
Bin Liang10
Estimated H-index: 10
(Kunming Institute of Zoology)
In order to understand the fundamental questions of the biology of life and to duplicate the pathogenesis of human diseases,animal models using different experimental animals,such as rodents,Drosophila,Caenorhabditis elegans,and zebrafish,have been established and used widely for many
2 Citations Source Cite
Published on Oct 1, 2015in Scientific Reports 4.12
Xianlong Wang6
Estimated H-index: 6
,
Jinwei Zhou1
Estimated H-index: 1
+ 11 AuthorsXiangnan Miao1
Estimated H-index: 1
Genetic engineering in livestock was greatly enhanced by the emergence of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9), which can be programmed with a single-guide RNA (sgRNA) to generate site-specific DNA breaks. However, the uncertainties caused by wide variations in sgRNA activity impede the utility of this system in generating genetically modified pigs. Here, we described a single blastocyst genotyping system to provide a simple and rapid solu...
37 Citations Source Cite
Published on Oct 1, 2015in International Journal of Pharmaceutics 3.86
Justin S. LaFountaine5
Estimated H-index: 5
(University of Texas at Austin),
Kristin Fathe6
Estimated H-index: 6
(University of Texas at Austin),
Hugh D. C. Smyth24
Estimated H-index: 24
In recent years, several new genome editing technologies have been developed. Of these the zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the CRISPR/Cas9 RNA-guided endonuclease system are the most widely described. Each of these technologies utilizes restriction enzymes to introduce a DNA double stranded break at a targeted location with the guide of homologous binding proteins or RNA. Such targeting is viewed as a significant advancement compared to...
38 Citations Source Cite
Published on Apr 1, 2016in Fungal Genetics and Biology 3.48
Mariana Schuster3
Estimated H-index: 3
(Max Planck Society),
Gabriel Schweizer6
Estimated H-index: 6
(Max Planck Society)
+ 1 AuthorsRegine Kahmann63
Estimated H-index: 63
(Max Planck Society)
Abstract This communication describes the establishment of the type II bacterial CRISPR–Cas9 system to efficiently disrupt target genes in the fungal maize pathogen Ustilago maydis . A single step transformation of a self-replicating plasmid constitutively expressing the U. maydis codon-optimized cas9 gene and a suitable sgRNA under control of the U. maydis U6 snRNA promoter was sufficient to induce genome editing. On average 70% of the progeny of a single transformant were disrupted within the ...
55 Citations Source Cite