{"title":"Efficient assembly of a synthetic attenuated SARS-CoV-2 genome in Saccharomyces cerevisiae using multi-copy yeast vectors","authors":"","doi":"10.1007/s12041-023-01455-5","DOIUrl":null,"url":null,"abstract":"<h3>Abstract</h3> <p><em>Saccharomyces cerevisiae</em> has been demonstrated to be an excellent platform for the multi-fragment assembly of large DNA constructs through its powerful homologous recombination ability. These assemblies have invariably used the stable centromeric single copy vectors. However, many applications of these assembled genomes would benefit from assembly in a higher copy number vector for improved downstream extraction of intact genomes from the yeast. A review of the literature revealed that large multi-fragment assemblies did not appear to have been attempted in multicopy vectors. Therefore, we devised a toolkit that would enable one to seamlessly transition with the same assembling fragments between a single copy and a multicopy vector. We evaluated the assembly of a 28 kb attenuated SARS-CoV-2 genome (lacking the N gene) from 10 fragments in both single copy and multicopy vector systems. Our results reveal that assembly was comparably efficient in the two vector systems. The findings should add to the synthetic biology toolkit of <em>S. cerevisiae</em> and should enable researchers to utilize any of these vector systems depending on their downstream applications.</p>","PeriodicalId":15907,"journal":{"name":"Journal of Genetics","volume":null,"pages":null},"PeriodicalIF":2.9000,"publicationDate":"2024-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Genetics","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1007/s12041-023-01455-5","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"EDUCATION & EDUCATIONAL RESEARCH","Score":null,"Total":0}
引用次数: 0
Abstract
Saccharomyces cerevisiae has been demonstrated to be an excellent platform for the multi-fragment assembly of large DNA constructs through its powerful homologous recombination ability. These assemblies have invariably used the stable centromeric single copy vectors. However, many applications of these assembled genomes would benefit from assembly in a higher copy number vector for improved downstream extraction of intact genomes from the yeast. A review of the literature revealed that large multi-fragment assemblies did not appear to have been attempted in multicopy vectors. Therefore, we devised a toolkit that would enable one to seamlessly transition with the same assembling fragments between a single copy and a multicopy vector. We evaluated the assembly of a 28 kb attenuated SARS-CoV-2 genome (lacking the N gene) from 10 fragments in both single copy and multicopy vector systems. Our results reveal that assembly was comparably efficient in the two vector systems. The findings should add to the synthetic biology toolkit of S. cerevisiae and should enable researchers to utilize any of these vector systems depending on their downstream applications.
期刊介绍：
The journal retains its traditional interest in evolutionary research that is of relevance to geneticists, even if this is not explicitly genetical in nature. The journal covers all areas of genetics and evolution,including molecular genetics and molecular evolution.It publishes papers and review articles on current topics, commentaries and essayson ideas and trends in genetics and evolutionary biology, historical developments, debates and book reviews. From 2010 onwards, the journal has published a special category of papers termed ‘Online Resources’. These are brief reports on the development and the routine use of molecular markers for assessing genetic variability within and among species. Also published are reports outlining pedagogical approaches in genetics teaching.
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