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| 1 | 遗传修饰技术在绵羊分子设计育种中的应用显示文摘利用遗传修饰技术可以使动物在遗传水平发生改变,实现在个体内表达外源基因或对其内源基因的功能造成影响。在动物育种中,可以利用遗传修饰技术在分子水平进行设计并实现品种的快速改良。从传统的遗传修饰技术、病毒载体、精子载体等介导的遗传修饰技术到新型人工核酸酶介导的基因编辑技术,尤其是CRISPR/Cas9为代表的人工核酸酶的运用使得基因编辑动物的制备变得更加高效快捷,并迅速在多个物种中得到应用。这些方法也已经拓展到了绵羊(Ovis aries)遗传育种领域。利用遗传修饰技术在绵羊中进行分子育种比传统的育种方式具有更大的优势,可以使用多种策略直接对性状进行快速改良,并且可以加快育种进程。本文详细介绍了遗传修饰技术在绵羊中的研究历程,探讨了通过遗传修饰技术进行绵羊分子设计育种的可能性,并提出以上技术和方法在绵羊育种中面临的问题和挑战。 | 王海涛 李亭亭 黄勋 马润林 刘秋月 | 2021 | 遗传2021，43，6： | 5 |
| 2 | Perspectives of pluripotent stem cells in livestock显示文摘The recent progress in derivation of pluripotent stem cells(PSCs)from farm animals opens new approaches not only for reproduction,genetic engineering,treatment and conservation of these species,but also for screening novel drugs for their efficacy and toxicity,and modelling of human diseases.Initial attempts to derive PSCs from the inner cell mass of blastocyst stages in farm animals were largely unsuccessful as either the cells survived for only a few passages,or lost their cellular potency;indicating that the protocols which allowed the derivation of murine or human embryonic stem(ES)cells were not sufficient to support the maintenance of ES cells from farm animals.This scenario changed by the innovation of induced pluripotency and by the development of the 3 inhibitor culture conditions to support naïve pluripotency in ES cells from livestock species.However,the long-term culture of livestock PSCs while maintaining the full pluripotency is still challenging,and requires further refinements.Here,we review the current achievements in the derivation of PSCs from farm animals,and discuss the potential application areas. | Dharmendra Kumar Thirumala R Talluri Naresh L Selokar Iqbal Hyder Wilfried A Ku | 2021 | World Journal of Stem Cells2021，13，1： | 4 |
| 3 | 单碱基编辑工具—腺嘌呤碱基编辑器ABE的研究进展显示文摘基因编辑技术CRISPR/Cas9自诞生以来,在基因敲除、敲入、碱基修复等领域得到了广泛的应用,但其效率低下,且存在非靶向切割,安全性较低,极大地限制了传统CRISPR/Cas9在高分辨率单碱基编辑中的应用。基于传统CRISPR/Cas9发展起来的碱基编辑器(base editor, BE)的出现则克服了这些弊端,其中,胞嘧啶碱基编辑器(cytosine base editor, CBE)能够使C·G转换为T·A,腺嘌呤碱基编辑器(adenine base editor, ABE)能够使A·T转换为G·C,都可以在不引入双链断裂的情况下实现高效的单碱基置换,这就避免了传统的CRISPR/Cas9在进行非同源末端连接(nonhomologous end-joining, NHEJ)时引入不可控的插入或缺失突变(Indels)。人类(Homo sapiens)大部分遗传疾病是由于碱基突变造成的,而单碱基编辑工具的出现能够在一定程度上纠正一定比例的致病性SNP,因此,其在动物模型构建、功能基因组学研究、分子育种、临床医学、转化医学等领域应用前景广阔。本文针对发展较晚、脱靶效率更低的腺嘌呤碱基编辑器ABE的原理、发展历程、应用和所面临的机遇与挑战进行了综述,以期为单碱基编辑技术的研究与应用提供参考。 | 李广栋 张鲁 富俊才 连正兴 刘国世 | 2019 | 农业生物技术学报2019，27，10： | 4 |
| 4 | Biological implications and limitations of a cynomolgus monkey with naturally occurring Parkinson's disease显示文摘We recently identified a cynomolgus monkey with naturally occurring Parkinson's disease(PD), indicating that PD may not be a uniquely human disease(Li et al., 2020). In our previous study, four lines of evidence, including typical PD clinical symptoms, pharmacological responses, pathological hallmarks, and genetic mutations, strongly supported the identification of a monkey with spontaneous PD(Figure 1). | Hao Li Yong-Gang Yao Xin-Tian Hu | 2021 | Zoological Research2021，42，2： | 4 |
| 5 | 利用单碱基编辑器定点突变猪肌肉生长抑制素基因的研究显示文摘【目的】利用单碱基编辑器在宁乡花猪肌肉生长抑制素(myostatin,MSTN)基因第2外显子处引入终止密码子,以获得MSTN基因表达沉默的肾成纤维细胞系,为后期培育MSTN碱基编辑猪奠定基础。【方法】首先在MSTN基因的第2外显子处设计1条单向导RNA(single guide RNA,sgRNA),将其连接至pMLM3636-puro质粒,形成重组表达载体pMLM3636-puro-MSTN,与含有红色荧光碱基编辑器YE1-BE3-FNLS共转入宁乡花猪肾成纤维细胞中,在红色荧光和嘌呤霉素双筛选条件下,挑取单克隆细胞,测序验证后,分析阳性单克隆细胞的蛋白表达情况。【结果】在MSTN基因第2外显子处发生了碱基定点突变,目标位点的氨基酸序列由色氨酸(TGG)转变成终止密码子(TAA),且G→A突变率为5.5%。Western blotting检测结果表明,试验组10号单克隆细胞的MSTN蛋白表达量与野生型相比降低了60%。【结论】本研究运用单碱基编辑技术在宁乡花猪MSTN基因编码区引入终止密码子,使翻译提前终止,导致蛋白表达量显著降低,为后期MSTN基因碱基编辑猪的生产奠定基础。 | 王晶 朱喆 张鹏 毕延震 | 2022 | 中国畜牧兽医2022，49，8： | 2 |
| 6 | CRISPR/Cas基因编辑技术治疗人类遗传性疾病的临床研究进展显示文摘随着人类基因组计划完成,人类遗传信息的解码推动着疾病诊疗迈入基因组学时代。许多医学领域中的“罕见病”和“不治之症”也被逐渐揭开了神秘的面纱,越来越多的疾病被确证为遗传性疾病。2012年,CRISPR/Cas(clustered regularly interspaced short palindromic repeats/CRISPR-associated enzyme)基因编辑技术的发现和应用,是近十年生物医学领域最具革命性的突破之一。该技术具有简便、高效、适用性广泛等特点,不但被广泛应用于基因功能研究,而且也在遗传病的基因治疗临床试验上取得重要进展。针对遗传背景明确的单基因遗传病,CRISPR/Cas可通过对目的DNA序列进行精准靶向编辑,实现对内源基因序列的改变或者基因功能的重新调控,为治愈部分重大遗传性疾病提供了新的工具和治疗策略。目前,多个基于CRISPR/Cas技术研发的基因药物已经进入临床试验,并取得了令人鼓舞的阶段性成果。本文就目前已公布临床试验的基因编辑药物研究进展进行回顾与展望。 | 胡思慧 刘倩宜 谢冬纯 黄军就 | 2022 | 生命科学2022，34，10： | 2 |
| 7 | Progress in in vitro culture and gene editing of porcine spermatogonial stem cells显示文摘Research on in vitro culture and gene editing of domestic spermatogonial stem cells (SSCs) is of considerable interest but remains a challenging issue in animal science. In recent years, some progress on the isolation, purification, and genetic manipulation of porcine SSCs has been reported. Here, we summarize the characteristics of porcine SSCs as well current advances in their in vitro culture, potential usage, and genetic manipulation. Furthermore, we discuss the current application of gene editing in pig cloning technology. Collectively, this commentary aims to summarize the progress made and obstacles encountered in porcine SSC research to better serve animal husbandry, improve livestock fecundity, and enhance potential clinical use. | Yi-Zhuo Sun Si-Tong Liu Xiao-Meng Li Kang Zou | 2019 | Zoological Research2019，40，5： | 1 |
| 8 | Double knock-in pig models with elements of binary Tet-On and phiC31 integrase systems for controllable and switchable gene expression显示文摘Inducible expression systems are indispensable for precise regulation and in-depth analysis of biological process.Binary Tet-On system has been widely employed to regulate transgenic expression by doxycycline.Previous pig models with tetracycline regulatory elements were generated through random integration.This process often resulted in uncertain expression and unpredictable phenotypes,thus hindering their applications.Here,by precise knock-in of binary Tet-On 3G elements into Rosa26 and Hipp11 locus,respectively,a double knock-in reporter pig model was generated.We characterized excellent properties of this system for controllable transgenic expression both in vitro and in vivo.Two att P sites were arranged to flank the td Tomato to switch reporter gene.Single or multiple gene replacement was efficiently and faithfully achieved in fetal fibroblasts and nuclear transfer embryos.To display the flexible application of this system,we generated a pig strain with Dox-inducing h KRASexpression through phiC31 integrase-mediated cassette exchange.After eight months of Dox administration,squamous cell carcinoma developed in the nose,mouth,and scrotum,which indicated this pig strain could serve as an ideal large animal model to study tumorigenesis.Overall,the established pig models with controllable and switchable transgene expression system will provide a facilitating platform for transgenic and biomedical research. | Qin Jin Xiaoyu Yang Shixue Gou Xiaoyi Liu Zhenpeng Zhuang Yanhui Liang Hui Shi Jiayuan Huang Han Wu Yu Zhao Zhen Ouyang Quanjun Zhang Zhaoming Liu Fangbing Chen Weikai Ge Jingke Xie Nan Li Chengdan Lai Xiaozhu Zhao Jiaowei Wang Meng Lian Lei Li Longquan Quan Yinghua Ye Liangxue Lai Kepin Wang | 2022 | Science China(Life Sciences)2022，65，11： | 1 |
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