"ATCGZP"六核酸分子合成生命遗传系统的设计与构建

"Artificially Expanded Genetic Information System" (AEGIS) is also known as "Expanded Genetic alphabets". Through modifying normal bases, it is designed to achieve the function of native nucleic acid with relatively independence. AEGIS has evolved since twenty-five years ago when this new concept was first introduced. Similar to native bases, plenty of research findings support that AEGIS are effective in replication, transcription, and even translation in vitro. However, no investigation is focus on the counterpart in vivo. Aiming to fill the gap, a living system that is capable of synthesizing, self-replicating and stably inheriting AEGIS in Escherichia coli is designed and created based on the achievements of the applicant's research into AEGIS. This work will begin a new chapter of AEGIS in vivo, which is not only an audacity of imagination of life engineering, but also a rewarding attempt in the field of synthesis of living systems from scratch. Moreover, it will greatly promote the development of "Chemical Synthetic Biology" and predictably break new ground in exploration of "Synthetic Biology".

人工扩展遗传信息系统（AEGIS）又称为人工扩展碱基字母，是一种通过对碱基的改造人工设计合成的可以行使天然核酸功能、而又具有相对独立性的人造DNA。人工扩展遗传信息系统发展至今已有25年的历史，在体外，已有足够多证据证明人造碱基可以行使和正常碱基一样的复制、转录甚至翻译的功能，但其体内研究始终没有开展。本项目正是针对这一空白，基于申请人在ATCGZP六分子人工扩展遗传信息系统领域所取得的一系列成果，以大肠杆菌为细胞底盘机架，设计并构建一种能实现ATCGZP六核酸分子体内合成、复制及稳定遗传的合成生命遗传系统。本项目的顺利实施为将人工扩展遗传信息系统由体外走向体内翻开了新的一页，这既是"生命工程化"的一次大胆设想，也是"从头合成生命系统"领域的一次有益的尝试，其也会对化学合成生物学研究领域的发展起到极大的推动促进作用，并可能为合成生物学开辟出一条新的、更安全的别样健康发展之路。

本研究以大肠杆菌为细胞底盘机架，设计并成功构建能够实现ATCGZP六核酸分子人工扩展遗传信息系统的体内合成、复制及遗传的大肠杆菌工程菌株。本工程菌通过外源导入果蝇脱氧核苷激酶的突变体Q81E-DmdNK，可高效实现外源dZ-OH和dP-OH的单磷酸转化（dZMP/dPMP）；本工程菌可利用E.coli内源的核苷单磷酸激酶（dNMK）和核苷二磷酸激酶（dNDK）实现dZMP/dPMP的多磷酸化内源合成相应三磷酸盐dZTP和dPTP。在成功实现dZTP/dPTP内源合成的基础上，本工程菌可进一步利用E.coli内源聚合酶实现了ATCGZP六核酸分子人工扩展遗传信息系统在大肠杆菌中的复制和遗传。当ATCGZP六核酸分子含有两个连续Z/P人工碱基对时,8小时后仍有近40% Z-P碱基对保有率。