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Updated: Feb 15, 2026
Hybrid De Novo Genome Assembly for the Generation of Complete Genomes of Urinary Bacteria using Short- and Long-read Sequencing Technologies
Published on: August 20, 2021
Efficient generation of complete sequences of MDR-encoding plasmids by rapid assembly of MinION barcoding sequencing
Ruichao Li1,2, Miaomiao Xie1, Ning Dong1
1Shenzhen Key Lab for Food Biological Safety Control, Food Safety and Technology Research Center, Hong Kong PolyU Shen Zhen Research Institute, Shenzhen, P. R. China.
Background:
Multidrug resistance (MDR)-encoding plasmids are considered major molecular vehicles responsible for transmission of antibiotic resistance genes among bacteria of the same or different species. Delineating the complete sequences of such plasmids could provide valuable insight into the evolution and transmission mechanisms underlying bacterial antibiotic resistance development. However, due to the presence of multiple repeats of mobile elements, complete sequencing of MDR plasmids remains technically complicated, expensive, and time-consuming.
Results:
Here, we demonstrate a rapid and efficient approach to obtaining multiple MDR plasmid sequences through the use of the MinION nanopore sequencing platform, which is incorporated in a portable device. By assembling the long sequencing reads generated by a single MinION run according to a rapid barcoding sequencing protocol, we obtained the complete sequences of 20 plasmids harbored by multiple bacterial strains. Importantly, single long reads covering a plasmid end-to-end were recorded, indicating that de novo assembly may be unnecessary if the single reads exhibit high accuracy.
Conclusions:
This workflow represents a convenient and cost-effective approach for systematic assessment of MDR plasmids responsible for treatment failure of bacterial infections, offering the opportunity to perform detailed molecular epidemiological studies to probe the evolutionary and transmission mechanisms of MDR-encoding elements.
Insights
This study presents a rapid, cost-effective method using nanopore sequencing to fully sequence multidrug resistance (MDR) plasmids. This approach aids in understanding antibiotic resistance gene transmission and evolution in bacteria.
Area of Science:
- Microbiology
- Genomics
- Molecular Biology
Background:
- Multidrug resistance (MDR)-encoding plasmids are key drivers of antibiotic resistance gene transmission between bacteria.
- Sequencing these plasmids is crucial for understanding resistance evolution but is often complex and costly.
- Existing methods face challenges due to repetitive elements within MDR plasmids.
Purpose of the Study:
- To develop a rapid and efficient method for obtaining complete sequences of MDR plasmids.
- To leverage portable nanopore sequencing technology for this purpose.
- To facilitate molecular epidemiological studies of antibiotic resistance.
Main Methods:
- Utilized the MinION nanopore sequencing platform with a rapid barcoding protocol.
- Generated long sequencing reads from multiple bacterial strains.
- Assembled long reads to obtain complete plasmid sequences.
Main Results:
- Successfully obtained complete sequences for 20 MDR plasmids.
- Demonstrated the ability to generate single, end-to-end plasmid reads.
- Highlighted the potential to bypass de novo assembly with high-accuracy long reads.
Conclusions:
- The developed workflow is convenient, cost-effective, and efficient for MDR plasmid sequencing.
- Enables systematic assessment of plasmids contributing to treatment failures.
- Facilitates detailed studies into the evolution and transmission of MDR elements.
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