Porcine Zygote Injection with Cas9/sgRNA Results in DMD-Modified Pig with Muscle Dystrophy
Abstract
:1. Introduction
2. Results
2.1. Validation of sgRNA Targeting DMD
2.2. Generation of DMD-Modified Pigs by Injecting Zygotes with Cas9/sgRNA
2.3. Genotype Analysis of DMD-Modified Pigs
2.4. Off-Target Detection
2.5. Phenotypic Analysis of the DMD-Modified Pig
3. Discussion
4. Materials and Methods
4.1. Animals
4.2. Construct and in Vitro Transcription
4.3. Pig Parthenogenetic Embryo Cas9/sgRNA Injection
4.4. Production of DMD-Modified Pigs by Injecting Zygotes with Cas9/sgRNA
4.5. T7EN1 Cleavage Assay and Sequencing
4.6. Off-Target Assay
4.7. Immunohistochemistry and Immunofluorescence
4.8. Hematoxylin and Eosin (H&E) Staining
4.9. Morphometric Analysis of Myofibers
5. Conclusion
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Yu, H.-H.; Zhao, H.; Qing, Y.-B.; Pan, W.-R.; Jia, B.-Y.; Zhao, H.-Y.; Huang, X.-X.; Wei, H.-J. Porcine Zygote Injection with Cas9/sgRNA Results in DMD-Modified Pig with Muscle Dystrophy. Int. J. Mol. Sci. 2016, 17, 1668. https://doi.org/10.3390/ijms17101668
Yu H-H, Zhao H, Qing Y-B, Pan W-R, Jia B-Y, Zhao H-Y, Huang X-X, Wei H-J. Porcine Zygote Injection with Cas9/sgRNA Results in DMD-Modified Pig with Muscle Dystrophy. International Journal of Molecular Sciences. 2016; 17(10):1668. https://doi.org/10.3390/ijms17101668
Chicago/Turabian StyleYu, Hong-Hao, Heng Zhao, Yu-Bo Qing, Wei-Rong Pan, Bao-Yu Jia, Hong-Ye Zhao, Xing-Xu Huang, and Hong-Jiang Wei. 2016. "Porcine Zygote Injection with Cas9/sgRNA Results in DMD-Modified Pig with Muscle Dystrophy" International Journal of Molecular Sciences 17, no. 10: 1668. https://doi.org/10.3390/ijms17101668