Development of rAAV-mediated method to modify common marmoset pre-implantation stage embryo genome

开发rAAV介导的方法来修改普通狨猴植入前阶段胚胎基因组

• 批准号: 23700521
• 负责人: OKADA Hironori
• 金额: $ 2.75万
• 依托单位: National Center of Neurology and Psychiatry
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Young Scientists (B)
• 财政年份: 2011
• 资助国家: 日本
• 起止时间: 2011 至 2013
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-23700521/
• 关键词: アデノ随伴ウイルスベクター; コモンマーモセット; 受精卵; 遺伝子挿入; 疾患モデル動物; アデノ随伴ウイルスベクタ ー

Although generation of transgenic marmosets with random insertion of lentivirus vector genome into embryo was reported, the phenotypes of such an animal would be diversified depending on the integration site. To establish more advanced method, adeno-associated virus vector (rAAV)-mediated transduction into marmoset embryo, and inclusion of several gene-editing systems into rAAV were examined. The results were as follows: 1) Enhancement of rAAV2 genome integration into a marmoset cell line due to addition of rAAV5 expressing AAV2Rep. 2) Construction of AAV vector plasmid carrying TALE with phi-Gamma catalytic domain. 3) Induction of indel mutation to HEK293 cells by transfection of AAV vector plasmid carrying CRISPR/Cas9 system, and production of rAAV from the plasmid. 4) Efficient rAAV9-mediated transduction into marmoset embryo, and consequent robust transgene expression in ICM. These resulting objects and findings would be useful for production of genome-edited non-human primate.

虽然已经报道了将慢病毒载体基因组随机插入胚胎的转基因狨猴，但这种动物的表型会因整合位点的不同而多样化。为了建立更先进的方法，研究了腺相关病毒载体（adeno-associated virus vector, rAAV）介导的狨猴胚胎转导，以及将几种基因编辑系统纳入rAAV。结果表明：1)通过添加表达AAV2Rep的rAAV5，增强了rAAV2基因组在狨猴细胞系中的整合。2)构建携带ph - γ催化结构域TALE的AAV载体质粒。3)用携带CRISPR/Cas9系统的AAV载体质粒转染HEK293细胞诱导indel突变，并从质粒中产生rAAV。4) raav9介导的高效绒猴胚胎转导，以及随后在ICM中的稳健转基因表达。这些结果和发现将对基因组编辑的非人类灵长类动物的生产有用。

项目成果

9型AAVベクターを用いた筋ジストロフィー犬胎仔への遺伝子導入と免疫寛容の誘導

使用9型AAV载体将基因转移至肌营养不良犬胎儿并诱导免疫耐受

• 发表时间: 2011
• 作者: Uechi M;Funayama M;Matsui Y;Mizuno T;Fujiwara M;Tajikawa T;Ohba K;Yamanami M;Watanabe T;Kanda K;Yaku H;Nakayama Y.;加藤天美;喜納裕美
• 通讯作者: 喜納裕美

Strategy for rAAV-mediated transduction of common marmoset skeletal muscle to generate NHP DMD model

rAAV 介导的普通狨猴骨骼肌转导生成 NHP DMD 模型的策略

• 发表时间: 2012
• 作者: Hironori Okada;Hidetoshi Ishibashi;Hiromi Hayashita-Kinoh;Tomoko Chiyo;Yuko Nitahara-Kasahara;Takashi Okada;and Shin'ichi Takeda
• 通讯作者: and Shin'ichi Takeda

rAAV8/9-Mediated Muscle Transduction with Tacrolimus in Non-Human Primate

rAAV8/9 介导的他克莫司在非人类灵长类动物中的肌肉转导

• 发表时间: 2012
• 作者: Akiko Ishii;Hironori Okada;Hiromi Hayashita-Kinou;Takashi Okada;Shin'ichi Takeda
• 通讯作者: Shin'ichi Takeda

rAAV9-Mediated Microdystrophin Gene Transfer with Immune Tolerance Induction Improves Dystrophic Phenotype of Canine X-Linked Muscular Dystrophy

rAAV9 介导的微小肌营养不良蛋白基因转移和免疫耐受诱导可改善犬 X 连锁肌营养不良症的营养不良表型

• 发表时间: 2012
• 作者: Hiromi Hayashita-Kinoh;et. al.
• 通讯作者: et. al.

Effective transgene expression in non-human primate muscle with AAV type 9 vectors following immune suppression

免疫抑制后，AAV 9 型载体在非人灵长类肌肉中有效表达转基因

• 发表时间: 2013
• 作者: Ishii A.;Okada H.;Hayashita-Kinoh H.;Shin J-H.;Okada T.;Takeda S.
• 通讯作者: Takeda S.