EAGER: DEVELOPMENT OF A TOOL CHAIN TO WRITE AND READ SELF-DOCUMENTING PLASMIDS

生命科学家在基础研究和新生物技术产品的开发中严重依赖工程DNA分子。这些分子的适当记录对于确保使用它们的过程的再现性和安全性至关重要。文档通常以标准计算机文件的形式出现。然而，在DNA分子和描述它们的文件之间没有直接的联系。跟踪DNA分子和描述它的计算机文件之间的关联证明是令人惊讶的挑战，因为DNA分子可以自行突变或在分子水平上被修改。同样，生物学家可以使用生物信息学软件编辑生物信息学文件。因此，在生命科学界（商业和研究）流通的分子的物理序列与其假定的参考序列之间存在广泛的差异。许多合成生物学家都有这样的经历，他们浪费了数周或数月的时间来研究质粒，结果发现它们与他们预期的不同。该项目将专注于使用计算机安全方法将质粒文档嵌入DNA分子本身。该项目支持培养一名化学工程博士后、一名计算机科学研究生和一名生物化学专业本科生。它还将发起一项社区努力，以评估自我记录质粒在保护合成生物学供应链方面的可能应用。它还将支持外联工作，通过新闻出版物和YouTube视频教育公众。软件将开源发布，质粒将存放在Addgene社区存储库中，数据将通过Figshare提供。该项目将开发DNAdoc，这是一种旨在确保生命科学研究中使用的质粒和其他合成DNA分子的身份和保真度的软件。DNAdoc将允许质粒和其他合成DNA的开发人员捕获质粒本身内的基本数据，并使用户能够从测序数据中检索这些信息。这些自我记录质粒将由两种类型的序列组成：1）功能序列，其编码基因和其他功能元件;以及2）记录序列，其唯一目的是编码关于功能序列的信息。将通过包括数字签名协议和纠错码在内的网络安全技术确保生物功能编码DNA序列与记录这些功能的序列之间结构联系的完整性。该项目包括两个目标。目标1的重点是编写自记录质粒和表征插入记录序列所产生的性能惩罚。目标2的重点是开发一个生物信息学工作流程，从Illumina测序数据中生成完整和高质量的序列文件。有关这项研究进展的信息可在https://peccoud.org上找到。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估来支持。

Life scientists rely heavily on engineered DNA molecules in basic research and the development of new biotechnology products. Proper documentation of these molecules is critical to ensure the reproducibility and safety of the processes in which they are used. Documentation generally comes in the form of standard computer files. However, there is no direct linkage between the DNA molecules and the files that describe them. Keeping track of the association between a DNA molecule and the computer file describing it proves surprisingly challenging because DNA molecules can mutate on their own or be modified at the molecular level. Similarly, biologists can edit bioinformatics files using bioinformatics software. As a result, there are widespread discrepancies between the physical sequences of the molecules in circulation in the life sciences community (both commercial and research) and their supposed reference sequence. Many synthetic biologists have the experience of wasting weeks or months working with plasmids that turned out to differ from what they were expecting. The project will focus on embedding the plasmid documentation in the DNA molecule itself using methods from computer security. The project supports the training of a postdoc in chemical engineering, a computer science graduate student, and an undergraduate student majoring in biochemistry. It will also initiate a community effort to evaluate the possible application of self-documenting plasmids to securing the synthetic biology supply chain. It will also support outreach efforts to educate the general public through news publications and YouTube videos. Software will be released open-source, plasmids will be deposited with Addgene a community repository, and data will be made available through Figshare.The project will develop DNAdoc, a software designed to ensure the identity and fidelity of plasmids and other synthetic DNA molecules used in life science research. DNAdoc will allow developers of plasmids and other synthetic DNA to capture essential data within the plasmid itself and enable users to retrieve this information from sequencing data. These self-documenting plasmids will be composed of two types of sequences 1) functional sequences, which encode genes and other functional elements and 2) documenting sequences whose only purpose is to encode information about the functional sequences. The integrity of the structural link between DNA sequences coding for biological functions and sequences documenting these functions will be ensured by cybersecurity techniques including digital signature protocols and error correction codes. The project includes two objectives. Objective 1 is focused on writing self-documenting plasmids and characterizing the performance penalty arising from the insertion of documenting sequences. The focus of Objective 2 is to develop a bioinformatics workflow producing a complete and high-quality sequence file from Illumina sequencing data. Information on the progress of this research will be found at https://peccoud.org .This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.