1. Single molecule sequencing of the M13 virus genome without amplification
- Author
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Michael W. Deem, Huan Jin, Li Gailing, Lidong Zeng, Jiankui He, Wu Ping, Huan Shang, Renli Zhang, Xu Weibin, Jinsen Cai, Pierre Ezanno, Zheng Jiao, Deng Liwei, Haomin Wu, Zhao Luyang, Yan Qin, Yan Li, Huan Zhao, Andrew X. Yang, and Zhou Zhiliang
- Subjects
0301 basic medicine ,Molecular biology ,lcsh:Medicine ,Artificial Gene Amplification and Extension ,Genome ,Polymerase Chain Reaction ,law.invention ,Database and Informatics Methods ,Sequencing techniques ,law ,Genomic library ,DNA sequencing ,Genome Sequencing ,lcsh:Science ,Polymerase chain reaction ,Base Composition ,Multidisciplinary ,High-Throughput Nucleotide Sequencing ,RNA sequencing ,Genomics ,Transcriptome Analysis ,Sequence Analysis ,Nucleic Acid Amplification Techniques ,Research Article ,Next-Generation Sequencing ,Bioinformatics ,Sequence alignment ,Computational biology ,Genome, Viral ,03 medical and health sciences ,Genomic Medicine ,Consensus sequence ,Genetics ,Biology and life sciences ,Base Sequence ,lcsh:R ,Computational Biology ,Nucleic acid amplification technique ,Vector Cloning ,Genome Analysis ,Genomic Libraries ,Research and analysis methods ,030104 developmental biology ,Molecular biology techniques ,lcsh:Q ,Sequence Alignment ,Cloning ,Bacteriophage M13 - Abstract
Next generation sequencing (NGS) has revolutionized life sciences research. However, GC bias and costly, time-intensive library preparation make NGS an ill fit for increasing sequencing demands in the clinic. A new class of third-generation sequencing platforms has arrived to meet this need, capable of directly measuring DNA and RNA sequences at the single-molecule level without amplification. Here, we use the new GenoCare single-molecule sequencing platform from Direct Genomics to sequence the genome of the M13 virus. Our platform detects single-molecule fluorescence by total internal reflection microscopy, with sequencing-by-synthesis chemistry. We sequenced the genome of M13 to a depth of 316x, with 100% coverage. We determined a consensus sequence accuracy of 100%. In contrast to GC bias inherent to NGS results, we demonstrated that our single-molecule sequencing method yields minimal GC bias.
- Published
- 2017