Rebustini Ivan T
Division of Genetics, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts.
Curr Protoc Cell Biol. 2017 Mar 3;74:19.19.1-19.19.17. doi: 10.1002/cpcb.15.
The increasing repertoire of microRNAs expressed during organ development and their role in regulating organ morphogenesis provide a compelling need to develop methods to assess microRNA function using various in vitro and in vivo experimental models. Methods to assess microRNA function during organ morphogenesis include transfection of microRNA inhibitors (antagomirs) and activators (mimics) into mouse embryonic explanted organs using liposomes, which can potentially result in low efficiency of transfection and off-target effects. We devised a method to assess microRNA function in explanted organs by transfecting antagomirs and mimics using peptide-based nanoparticles, increasing functional microRNA targeting efficiency, and decreasing off-target effects. Our method can be applied to a variety of embryonic organs that can be explanted and provides an alternative to efficiently and functionally prioritize microRNAs during organ morphogenesis for further in vivo genetic approaches. © 2017 by John Wiley & Sons, Inc.
在器官发育过程中表达的微小RNA种类不断增加,及其在调节器官形态发生中的作用,迫切需要开发利用各种体外和体内实验模型来评估微小RNA功能的方法。评估器官形态发生过程中微小RNA功能的方法包括使用脂质体将微小RNA抑制剂(抗反义寡核苷酸)和激活剂(模拟物)转染到小鼠胚胎外植器官中,这可能会导致转染效率低和脱靶效应。我们设计了一种方法,通过使用基于肽的纳米颗粒转染抗反义寡核苷酸和模拟物来评估外植器官中的微小RNA功能,提高功能性微小RNA靶向效率,并减少脱靶效应。我们的方法可应用于多种可外植的胚胎器官,并为在器官形态发生过程中高效且功能性地对微小RNA进行优先级排序提供了一种替代方法,以便进一步进行体内遗传学研究。© 2017 约翰威立父子出版公司