{"title":"测序平台对使用 Hawk-Seq™ 检测化学变异灵敏度的影响。","authors":"Sayaka Hosoi, Takako Hirose, Shoji Matsumura, Yuki Otsubo, Kazutoshi Saito, Masaaki Miyazawa, Takayoshi Suzuki, Kenichi Masumura, Kei-Ichi Sugiyama","doi":"10.1186/s41021-024-00313-9","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Error-corrected next-generation sequencing (ecNGS) technologies have enabled the direct evaluation of genome-wide mutations after exposure to mutagens. Previously, we reported an ecNGS methodology, Hawk-Seq™, and demonstrated its utility in evaluating mutagenicity. The evaluation of technical transferability is essential to further evaluate the reliability of ecNGS-based assays. However, cutting-edge sequencing platforms are continually evolving, which can affect the sensitivity of ecNGS. Therefore, the effect of differences in sequencing instruments on mutation data quality should be evaluated.</p><p><strong>Results: </strong>We assessed the performance of four sequencing platforms (HiSeq2500, NovaSeq6000, NextSeq2000, and DNBSEQ-G400) with the Hawk-Seq™ protocol for mutagenicity evaluation using DNA samples from mouse bone marrow exposed to benzo[a]pyrene (BP). The overall mutation (OM) frequencies per 10<sup>6</sup> bp in vehicle-treated samples were 0.22, 0.36, 0.46, and 0.26 for HiSeq2500, NovaSeq6000, NextSeq2000, and DNBSEQ-G400, respectively. The OM frequency of NextSeq2000 was significantly higher than that of HiSeq2500, suggesting the difference to be based on the platform. The relatively higher value in NextSeq2000 was a consequence of the G:C to C:G mutations in NextSeq2000 data (0.67 per 10<sup>6</sup> G:C bp), which was higher than the mean of the four platforms by a ca. of 0.25 per 10<sup>6</sup> G:C bp. A clear dose-dependent increase in G:C to T:A mutation frequencies was observed in all four sequencing platforms after BP exposure. The cosine similarity values of the 96-dimensional trinucleotide mutation patterns between HiSeq and the three other platforms were 0.93, 0.95, and 0.92 for NovaSeq, NextSeq, and DNBSeq, respectively. These results suggest that all platforms can provide equivalent data that reflect the characteristics of the mutagens.</p><p><strong>Conclusions: </strong>All platforms sensitively detected mutagen-induced mutations using the Hawk-Seq™ analysis. The substitution types and frequencies of the background errors differed depending on the platform. The effects of sequencing platforms on mutagenicity evaluation should be assessed before experimentation.</p>","PeriodicalId":12709,"journal":{"name":"Genes and Environment","volume":"46 1","pages":"20"},"PeriodicalIF":2.7000,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11462924/pdf/","citationCount":"0","resultStr":"{\"title\":\"Effect of sequencing platforms on the sensitivity of chemical mutation detection using Hawk-Seq™.\",\"authors\":\"Sayaka Hosoi, Takako Hirose, Shoji Matsumura, Yuki Otsubo, Kazutoshi Saito, Masaaki Miyazawa, Takayoshi Suzuki, Kenichi Masumura, Kei-Ichi Sugiyama\",\"doi\":\"10.1186/s41021-024-00313-9\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Error-corrected next-generation sequencing (ecNGS) technologies have enabled the direct evaluation of genome-wide mutations after exposure to mutagens. Previously, we reported an ecNGS methodology, Hawk-Seq™, and demonstrated its utility in evaluating mutagenicity. The evaluation of technical transferability is essential to further evaluate the reliability of ecNGS-based assays. However, cutting-edge sequencing platforms are continually evolving, which can affect the sensitivity of ecNGS. Therefore, the effect of differences in sequencing instruments on mutation data quality should be evaluated.</p><p><strong>Results: </strong>We assessed the performance of four sequencing platforms (HiSeq2500, NovaSeq6000, NextSeq2000, and DNBSEQ-G400) with the Hawk-Seq™ protocol for mutagenicity evaluation using DNA samples from mouse bone marrow exposed to benzo[a]pyrene (BP). The overall mutation (OM) frequencies per 10<sup>6</sup> bp in vehicle-treated samples were 0.22, 0.36, 0.46, and 0.26 for HiSeq2500, NovaSeq6000, NextSeq2000, and DNBSEQ-G400, respectively. The OM frequency of NextSeq2000 was significantly higher than that of HiSeq2500, suggesting the difference to be based on the platform. The relatively higher value in NextSeq2000 was a consequence of the G:C to C:G mutations in NextSeq2000 data (0.67 per 10<sup>6</sup> G:C bp), which was higher than the mean of the four platforms by a ca. of 0.25 per 10<sup>6</sup> G:C bp. A clear dose-dependent increase in G:C to T:A mutation frequencies was observed in all four sequencing platforms after BP exposure. The cosine similarity values of the 96-dimensional trinucleotide mutation patterns between HiSeq and the three other platforms were 0.93, 0.95, and 0.92 for NovaSeq, NextSeq, and DNBSeq, respectively. These results suggest that all platforms can provide equivalent data that reflect the characteristics of the mutagens.</p><p><strong>Conclusions: </strong>All platforms sensitively detected mutagen-induced mutations using the Hawk-Seq™ analysis. The substitution types and frequencies of the background errors differed depending on the platform. The effects of sequencing platforms on mutagenicity evaluation should be assessed before experimentation.</p>\",\"PeriodicalId\":12709,\"journal\":{\"name\":\"Genes and Environment\",\"volume\":\"46 1\",\"pages\":\"20\"},\"PeriodicalIF\":2.7000,\"publicationDate\":\"2024-10-09\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11462924/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Genes and Environment\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1186/s41021-024-00313-9\",\"RegionNum\":4,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"GENETICS & HEREDITY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Genes and Environment","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1186/s41021-024-00313-9","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"GENETICS & HEREDITY","Score":null,"Total":0}
Effect of sequencing platforms on the sensitivity of chemical mutation detection using Hawk-Seq™.
Background: Error-corrected next-generation sequencing (ecNGS) technologies have enabled the direct evaluation of genome-wide mutations after exposure to mutagens. Previously, we reported an ecNGS methodology, Hawk-Seq™, and demonstrated its utility in evaluating mutagenicity. The evaluation of technical transferability is essential to further evaluate the reliability of ecNGS-based assays. However, cutting-edge sequencing platforms are continually evolving, which can affect the sensitivity of ecNGS. Therefore, the effect of differences in sequencing instruments on mutation data quality should be evaluated.
Results: We assessed the performance of four sequencing platforms (HiSeq2500, NovaSeq6000, NextSeq2000, and DNBSEQ-G400) with the Hawk-Seq™ protocol for mutagenicity evaluation using DNA samples from mouse bone marrow exposed to benzo[a]pyrene (BP). The overall mutation (OM) frequencies per 106 bp in vehicle-treated samples were 0.22, 0.36, 0.46, and 0.26 for HiSeq2500, NovaSeq6000, NextSeq2000, and DNBSEQ-G400, respectively. The OM frequency of NextSeq2000 was significantly higher than that of HiSeq2500, suggesting the difference to be based on the platform. The relatively higher value in NextSeq2000 was a consequence of the G:C to C:G mutations in NextSeq2000 data (0.67 per 106 G:C bp), which was higher than the mean of the four platforms by a ca. of 0.25 per 106 G:C bp. A clear dose-dependent increase in G:C to T:A mutation frequencies was observed in all four sequencing platforms after BP exposure. The cosine similarity values of the 96-dimensional trinucleotide mutation patterns between HiSeq and the three other platforms were 0.93, 0.95, and 0.92 for NovaSeq, NextSeq, and DNBSeq, respectively. These results suggest that all platforms can provide equivalent data that reflect the characteristics of the mutagens.
Conclusions: All platforms sensitively detected mutagen-induced mutations using the Hawk-Seq™ analysis. The substitution types and frequencies of the background errors differed depending on the platform. The effects of sequencing platforms on mutagenicity evaluation should be assessed before experimentation.
期刊介绍：
Genes and Environment is an open access, peer-reviewed journal that aims to accelerate communications among global scientists working in the field of genes and environment. The journal publishes articles across a broad range of topics including environmental mutagenesis and carcinogenesis, environmental genomics and epigenetics, molecular epidemiology, genetic toxicology and regulatory sciences.
Topics published in the journal include, but are not limited to, mutagenesis and anti-mutagenesis in bacteria; genotoxicity in mammalian somatic cells; genotoxicity in germ cells; replication and repair; DNA damage; metabolic activation and inactivation; water and air pollution; ROS, NO and photoactivation; pharmaceuticals and anticancer agents; radiation; endocrine disrupters; indirect mutagenesis; threshold; new techniques for environmental mutagenesis studies; DNA methylation (enzymatic); structure activity relationship; chemoprevention of cancer; regulatory science. Genetic toxicology including risk evaluation for human health, validation studies on testing methods and subjects of guidelines for regulation of chemicals are also within its scope.
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