关键词: 自动化液体处理, 交叉污染, 全基因组序列测定, 病原体网络化监测

Abstract: Objective To establish a high-throughput semi-automatic bacterial whole-genome sequencing system to meet the needs of bacterial pathogen surveillance based on the genome sequence. Methods An automated liquid handler, disposable mechanical pipette tips, a 96-well nucleic acid extraction kit, an automated sequencing library construction system, and a sequencer were used to realize the semi-automation of bacterial genome sequencing. Results The distance between the mechanical pipette tip and the bottom of the sample well, the mixing speed of liquid, and the volume ratio of the mixed liquid to the sample each time were closely related to the cross-contamination of the samples. When the distance between the mechanical pipette tip and the bottom of the sample well was 2 mm, the mixing speed of liquid was 15% of the full speed, and the volume ratio of the mixed liquid to the sample was 50% each time, the contamination rate was 0. Compared with the manual procedure, this high-throughput semi-automatic bacterial whole-genome sequencing system improved the work efficiency by 4-fold. Conclusion A high-throughput semi-automatic bacterial whole-genome sequencing system is established, which can meet the needs of bacterial pathogen surveillance based on the genome sequence.

Key words: Automated liquid handling, Cross-contamination, Whole-genome sequencing, Pathogen surveillance network