A DNA Part Library for Reliable Engineering of the Emerging Model Nematode Symbiotic Bacterium Xenorhabdus griffiniae HGB2511.

IF 3.9 2区生物学 Q1 BIOCHEMICAL RESEARCH METHODS

ACS Synthetic Biology Pub Date : 2025-09-25 DOI:10.1021/acssynbio.5c00414

Elin M Larsson, Olivia Y Wang, Richard M Murray

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引用次数: 0

Abstract

Xenorhabdus griffiniae is a bacterium that lives inside the intestine of the entomopathogenic nematode Steinernema hermaphroditum and partners with the nematode to infect and kill insect larvae in soil. The construction of gene circuits, such as reporters, in X. griffiniae would provide tools to study and better understand the symbiotic relationship it has with its host. However, because X. griffiniae is not a model organism, information about gene circuit construction in X. griffiniae is limited. We developed and characterized a DNA part library similar to the CIDAR MoClo extension library for E. coli to allow more efficient construction of genetic circuits in X. griffiniae. TurboRFP expressing strains with different constitutive Anderson promoters and different ribosome binding sites (RBS) were constructed to quantify promoter and RBS strengths in X. griffiniae. Furthermore, two fluorescent proteins sfGFP and sfYFP as well as the bioluminescent luxCDABE operon were added to the part library and successfully expressed in X. griffiniae. We then used the characterized parts of the cell to build and characterize IPTG inducible constructs.

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新兴模式线虫共生细菌格里菲尼异芽胞杆菌HGB2511的可靠工程DNA片段库

griffiniae Xenorhabdus是一种生活在昆虫病原线虫雌雄斯坦线虫（Steinernema hermaphroditum）肠道内的细菌，与该线虫合作感染并杀死土壤中的昆虫幼虫。在X. griffiniae中构建基因回路，如报告基因，将为研究和更好地理解其与宿主的共生关系提供工具。然而，由于griffiniae不是模式生物，关于griffiniae基因回路构建的信息有限。我们开发并鉴定了一个类似于大肠杆菌CIDAR MoClo扩展库的DNA片段库，以便更有效地构建X. griffiniae的遗传电路。构建具有不同组成型Anderson启动子和不同核糖体结合位点（RBS）的TurboRFP表达菌株，定量测定启动子和RBS在X. griffiniae中的强度。在部分文库中添加了两个荧光蛋白sfGFP和sfYFP以及生物发光的luxCDABE操纵子，并在X. griffiniae中成功表达。然后，我们使用细胞的特征部分来构建和表征IPTG诱导结构。

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来源期刊

ACS Synthetic Biology 生物-

CiteScore

8.00

自引率

10.60%

发文量

380

审稿时长

6-12 weeks

期刊介绍： The journal is particularly interested in studies on the design and synthesis of new genetic circuits and gene products; computational methods in the design of systems; and integrative applied approaches to understanding disease and metabolism. Topics may include, but are not limited to: Design and optimization of genetic systems Genetic circuit design and their principles for their organization into programs Computational methods to aid the design of genetic systems Experimental methods to quantify genetic parts, circuits, and metabolic fluxes Genetic parts libraries: their creation, analysis, and ontological representation Protein engineering including computational design Metabolic engineering and cellular manufacturing, including biomass conversion Natural product access, engineering, and production Creative and innovative applications of cellular programming Medical applications, tissue engineering, and the programming of therapeutic cells Minimal cell design and construction Genomics and genome replacement strategies Viral engineering Automated and robotic assembly platforms for synthetic biology DNA synthesis methodologies Metagenomics and synthetic metagenomic analysis Bioinformatics applied to gene discovery, chemoinformatics, and pathway construction Gene optimization Methods for genome-scale measurements of transcription and metabolomics Systems biology and methods to integrate multiple data sources in vitro and cell-free synthetic biology and molecular programming Nucleic acid engineering.