[3S05Aa] 21. Open Topics in Biotechnology - Part 1

3:45 PM - 4:00 PM JST(6:45 AM - 7:00 AM UTC)

[3EMT-01-OP] Pipeline and Insulated Dual-Reporter System to Predict and Quantify Strong Terminators from RNA-Seq Datasets for Metabolic Engineering

○Yunhao Chen¹, Qiaoning He¹, Qiushi Ye¹, Jiapeng Li¹, Changzheng Li¹, Shihui Yang¹ (1. Hubei Univ. (China))

Keywords:

Terminator,RNA-seq,Insulated dual-reporter system,Termination efficiency,Cross-species compatibility,Synthetic biology

The limited availability of genetic regulatory parts, particularly terminators of non-model microorganisms, poses a challenge on precise gene regulation to construct effective and efficient microbial cell factories. In this study, an Insulated dual-reporter system (IDRS) was developed to quantify the termination efficiency of candidate terminators accurately, and an IDRS-RNA-Seq pipeline for strong terminator prediction and quantification was further established combining RNA-seq datasets and the IDRS. RNA-seq data from Zymomonas mobilis and Escherichia coli were used to predict strong terminators with 48 and 41 out of all 48 and 272 strong terminator candidates of Z. mobilis and E. coli quantified using the IDRS, respectively. The positive rate for strong terminator prediction in Z. mobilis was 94% and 71% in E. coli. Our findings also demonstrated robust cross-species compatibility of strong terminators in Z. mobilis and E. coli, and the upstream sequence length and temperature influenced termination efficiency. This research provides a generalizable framework for high-throughput prediction, quantification and cross-species compatibility analysis of strong bacterial terminators genome-widely, thus enriching the biological parts of prokaryotic terminator libraries to accelerate the development of efficient and robust microbial cell factories with stable metabolic pathway and precise transcriptional control.