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4 results on '"Haiwen Zhong"'

1. Efficient and Safe Editing of Porcine Endogenous Retrovirus Genomes by Multiple-Site Base-Editing Editor

Author

Shuwen Zheng, Haiwen Zhong, Xiaoqing Zhou, Min Chen, Wansheng Li, Yin Zi, Yue Chi, Jinling Wang, Wei Zheng, Qingjian Zou, Liangxue Lai, and Chengcheng Tang

Subjects
porcine endogenous retroviruses, base editors, multiple loci, Cytology, QH573-671
Abstract

Gene-modified miniature pigs serve as alternative tissue and organ donors for xenotransplantation to alleviate the shortage of human allogenic organs. However, the high copy number of porcine endogenous retrovirus (PERV) genomes integrates with the porcine genome, which has a potential risk of cross-species transmission and hinders the clinical practice of xenotransplantation. Recently, CRISPR/Cas9 has been used to inactivate PERVs. However, Cas9 also triggers severe DNA damage at multiple integrated PERV sites in the porcine genome, which induces senescence and apoptosis of porcine cells. In this study, the cytosine base editor (CBE), an efficient and safe editor that does not cause DNA double strand breaks (DSBs), was used for PERV editing to reduce cytotoxic effects. Seven sgRNAs were set to target gag and pol loci of PERVs to induce premature stop codons. We found that approximately 10% of cell clones were completely inactivated for PERVs in pig ST cells, and the plasmid that was used for editing the PERVs did not integrate into host genome and influence the karyotype of the modified cells. Our studies offer a powerful and safe strategy for further generating PERV-knockout pigs using base editors.

Published
2022
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2. Pig Coat Color Manipulation by MC1R Gene Editing

Author

Haiwen Zhong, Jian Zhang, Cheng Tan, Junsong Shi, Jie Yang, Gengyuan Cai, Zhenfang Wu, and Huaqiang Yang

Subjects
coat color, CRISPR, gene editing, homologous recombination, MC1R, pig, Biology (General), QH301-705.5, Chemistry, QD1-999
Abstract

Black coat color in pigs is determined by the dominant E allele at the MC1R locus. Through comparing MC1R gene sequences between recessive e and dominant ED1 alleles, we identified four missense mutations that could affect MC1R protein function for eumelanin synthesis. With the aim of devising a genetic modification method for pig coat color manipulation, we mutated the e allele in the Duroc breed to the dominant ED1 allele using CRISPR-mediated homologous recombination for the four mutation substitutions at the MC1R locus. The MC1R-modified Duroc pigs generated using the allele replacement strategy displayed uniform black coat color across the body. A genotyping assay showed that the MC1R-modified Duroc pigs had a heterozygous ED1/e allele at the MC1R locus; in addition, the pigs remained in the Duroc genetic background. Our work offers a gene editing method for pig coat color manipulation, which could value the culture of new pig varieties meeting the needs of diversified market.

Published
2022
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3. Research on Official Media Coverage and Communication Mode on Different SNS Platforms-Taking People's Daily's Report on Tangshan Restaurant Attack as an Example

Author

Yulin An, Shifang Cai, Haiwen Zhong, and Ziying Zhou

Abstract

In the era of media convergence, there are more diverse forms of communication in official media. Therefore, people can access the latest news more easily and quickly through new media. This study uses the methods of content analysis and frame analysis to investigate the reporting framework mode adopted by People's Daily on different platforms in social emergencies and whether the mainstream reporting mode of the same platform will change with the change of timeline, so as to explore how official media adapt to the trend of media convergence and compete for the field of public opinion through the differentiation of official discourse space. The results are as below: Firstly, People's Daily's TikTok account mainly adopts episodic framing, while other platforms and accounts mainly adopt thematic framing. Therefore, People's Daily's reporting strategy is mainly based on thematic framing; Secondly, Traditional official media have begun to break through the restrictions of official discourse space and are trying to adjust from within, actively accepting unofficial discourse space and allowing the emergence of different voices and diversified expressions.

Published
2023

4. Establishment of a pig CRISPR/Cas9 knockout library for functional gene screening in pig cells

Author

Guoling Li, Xiaohui Yang, Chuanzhao Yu, Haiwen Zhong, Huaqiang Yang, and Zhenfang Wu

Subjects
Gene Editing, Swine, Cas9, Lentivirus, Diphthamide, General Medicine, Computational biology, Biology, Endonucleases, Applied Microbiology and Biotechnology, Genome, Phenotype, Gene Knockout Techniques, chemistry.chemical_compound, chemistry, Animals, Molecular Medicine, CRISPR, CRISPR-Cas Systems, DPH1, Gene, Gene knockout, Gene Library, RNA, Guide, Kinetoplastida
Abstract

Background As an important farm animal, pig functional genomic study can help understand the molecular mechanism related to the key economic traits of pig, such as growth, reproduction, or disease. The genome-scale library based on clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated endonuclease Cas9 (Cas9) system facilitates discovery of key genes involved in a specific function or phenotype, allowing for an effective "phenotype-to-genotype" strategy for functional genomic study. Methods and results We designed and constructed a pig genome-scale CRISPR/Cas9 knockout library targeting 16888 genes with 970001 unique sgRNAs. The library is a single-vector system including both Cas9 and sgRNA, and packaged into lentiviral vectors for an easy cell delivery for screening. To establish a screening method in pig cells, we used diphtheria toxin (DT)-induced cell death as a model to screen the host genes critical for DT toxicity in pig PK-15 cells. After lentiviral transduction and two sequential screening with DT treatment, the highest-ranking candidates we identified were previously validated genes, HBEGF, DPH1, DPH2, DPH3, DPH5, DNAJC24, and ZBTB17, which are DT receptor and the key factors involved in biosynthesis of diphthamide, the target of DT action. The function and gene essentiality of candidates were further confirmed by gene knockout and DT toxicity assay in PK-15 cells. Conclusions Our CRISPR knockout library targeting pig whole genome establishes a promising platform for pig functional genomic analysis. This article is protected by copyright. All rights reserved.

Published
2021

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