Amneh Aoudi, Ossama Labiad, Ramdane Igalouzene, Ousséma Mejri, Maxime Sanchez, Maxime Sanchez
{"title":"Mobilization of Plasmids From Bacteria Into Diatoms by Conjugation Technique","authors":"Amneh Aoudi, Ossama Labiad, Ramdane Igalouzene, Ousséma Mejri, Maxime Sanchez, Maxime Sanchez","doi":"10.21769/BioProtoc.4945","DOIUrl":null,"url":null,"abstract":"Diatoms serve as a source for a variety of compounds with particular biotechnological interest. Therefore, redirecting the flow to a specific pathway requires the elucidation of the gene’s specific function. The most commonly used method in diatoms is biolistic transformation, which is a very expensive and time-consuming method. The use of episomes that are maintained as closed circles at a copy number equivalent to native chromosomes has become a useful genetic system for protein expression that avoids multiple insertions, position-specific effects on expression, and potential knockout of non-targeted genes. These episomes can be introduced from bacteria into diatoms via conjugation. Here, we describe a detailed protocol for gene expression that includes 1) the gateway cloning strategy and 2) the conjugation protocol for the mobilization of plasmids from bacteria to diatoms.","PeriodicalId":93907,"journal":{"name":"Bio-protocol","volume":null,"pages":null},"PeriodicalIF":1.0000,"publicationDate":"2024-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Bio-protocol","FirstCategoryId":"0","ListUrlMain":"https://doi.org/10.21769/BioProtoc.4945","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Diatoms serve as a source for a variety of compounds with particular biotechnological interest. Therefore, redirecting the flow to a specific pathway requires the elucidation of the gene’s specific function. The most commonly used method in diatoms is biolistic transformation, which is a very expensive and time-consuming method. The use of episomes that are maintained as closed circles at a copy number equivalent to native chromosomes has become a useful genetic system for protein expression that avoids multiple insertions, position-specific effects on expression, and potential knockout of non-targeted genes. These episomes can be introduced from bacteria into diatoms via conjugation. Here, we describe a detailed protocol for gene expression that includes 1) the gateway cloning strategy and 2) the conjugation protocol for the mobilization of plasmids from bacteria to diatoms.