基于TALENs系统的食品微生物新型基因组编辑育种技术的建立

TALENs system is a new technique which can be used for Genome editing in vivo. TALENs is much more precise, effective and convenient than old methods and was named "Method of the Year 2011" by Nature. With the rapid development of Genomics, more and more strains' genome were sequenced. The traditional gene targeting method is out-of-date for its high off-targeting rate and inefficiency. Targeting accuracy requirements are higher for food microorganisms, for the consideration of food safety. TALENs is an artificial genetic tool which can be designed to target any desired loci on DNA sequences precisely and induce DNA double-strand breaks. Now, there are no reports about the application of TALENs in the Breeding of food microbes. In order to apply TALENs for any special food microorganism, foundational research needed to be done first to fine-tune the system . We choose Aspergillus niger which was used in food industry as the model system to study, intend to use TALENs to edit its genome by inserting or deleting DNA segments precisely, effectively and conveniently. We plan to establish a new method for microbial breeding and a protocol to assay and evaluate it which will be a new technique platform for further study.

TALENs系统是近期出现的一种对生物体基因组的遗传信息进行编辑改造的新技术，它比传统的基因打靶技术更为精确、高效、简便，并被Nature评为2011年的年度技术。在基因组学快速发展的时代背景下，大量微生物基因组被破译，而已经使用近30年的传统基因打靶技术，因脱靶率较高、工作效率低等诸多不足，已经渐渐不能满足微生物育种的发展。对于食品微生物而言，出于安全性考虑，对打靶精确度要求更高。TALENs作为一种人造的分子工具，它可以被设计成与任意的DNA序列结合并切割DNA分子，潜力巨大。目前，不仅尚未有关于TALENs系统在食品微生物育种领域应用的报道，而且由于微生物细胞的特殊性，其应用也需要进行探索性的基础研究。本研究拟选取食品行业常用的黑曲霉作为研究模型，综合使用生物信息学、分子生物学和微生物遗传学等技术手段，尝试在其基因组上使用TALENs系统精确地、简便和高效地插入与敲除DNA片段。

本研究成功构建了一套针对黑曲霉细胞的基因编辑工具，实现了对黑曲霉基因组的编辑，并分析和评价了其编辑效果，本研究还成功探索出一套全新的针对黑曲霉的外源核酸转化体系，提高了将外源核酸导入细胞的效率。