基于“人造精子细胞”的基因组标签计划

“Artificial spermatid”-based Genome tagging project

蛋白质是一切生命活动的基础,蛋白质研究特别是生命活动中蛋白质功能网络调控的揭示成为继人类基因组计划后广受关注的重要科学问题.然而,大规模开展蛋白质研究缺乏一个高效、标准的平台,构建基因组范围的蛋白质标签模式生物资源平台是解决这一问题的关键.虽然在酵母、线虫、果蝇等模式生物上已经实现了基因组范围的蛋白质标签文库的构建,但是在小鼠个体水平上,基因组范围的蛋白质标签的构建仍然困难重重.基因组标签计划(Genome tagging project, GTP)的提出是基于孤雄单倍体胚胎干细胞(又称为"人造精子细胞")介导的半克隆技术和CRISPR-Cas9基因编辑技术的发展:首先在"人造精子细胞"上利用CRISPR-Cas9技术对特定编码蛋白质基因进行标签序列的原位插入获得相应的细胞系,进一步通过卵子注射即能获得携带标签的小鼠模型.因此,该策略简单、高效、成本低,可以用于基因组范围制备蛋白质标签小鼠. GTP的目标是给小鼠中的每个蛋白质带上标签, GTP的实施有望实现以一套标准化的方法来对每个蛋白质进行在体研究,将促进生命科学的发展.

Protein is the basis of all biological processes. Protein studies, especially the discovery of protein functional regulation network in life activities, have become an important scientific issue that has drawn increasing attention from scientists after the Human Genome Project. However, large-scale protein research needs an efficient and standard platform. Building a resource platform of model organism where every protein is tagged at organismal level is the key to solving this problem. However, although genome-wide protein labelling has been achieved in Saccharomyces cerevisiae, Caenorhabditis elegans and Drosophila, genome-wide protein tagging in mice is still very challenging. Genome tagging project(GTP) in mice has been proposed based on two state-of-art biotechnologies, i.e., androgenetic haploid embryonic stem cell(so-called "artificial spermatid")-mediated semi-cloning technology and CRISPR-Cas9-based genome-editing technology. GTP involves two steps: first, the "artificial spermatid" lines carrying a tagged protein are constructed in vitro by endogenously inserting a tag sequence into a specific protein-coding gene via CRISPR-Cas9;second, the tagged "artificial spermatids" can be injected into oocytes to obtain tagged mice in one step. This strategy by combining"artificial spermatids" and CRISPR-Cas9 is simple, efficient, and low-cost, and thus can be used to generate tagged mice at the genome-wide scale. The goal of GTP is to tag every protein in mice and, we believe, the implementation of GTP will promote to investigate the protein expression, localization and function in vivo through standardized tag antibody-based assays, which will greatly enhance the advances of life sciences.