Buck Jochen M, Río Bártulos Carolina, Gruber Ansgar, Kroth Peter G
Department of Biology, University of Konstanz, Konstanz, Germany.
Institute of Parasitology, Biology Centre, Czech Academy of Sciences, České Budějovice, Czech Republic.
PeerJ. 2018 Nov 14;6:e5884. doi: 10.7717/peerj.5884. eCollection 2018.
Most genetic transformation protocols for the model diatom rely on one of two available antibiotics as selection markers: Zeocin (a formulation of phleomycin D1) or nourseothricin. This limits the number of possible consecutive genetic transformations that can be performed. In order to expand the biotechnological possibilities for , we searched for additional antibiotics and corresponding resistance genes that might be suitable for use with this diatom. Among the three different antibiotics tested in this study, blasticidin-S and tunicamycin turned out to be lethal to wild-type cells at low concentrations, while voriconazole had no detectable effect on . Testing the respective resistance genes, we found that the blasticidin-S deaminase gene () effectively conferred resistance against blasticidin-S to . Furthermore, we could show that expression of did not lead to cross-resistances against Zeocin or nourseothricin, and that genetically transformed cell lines with resistance against Zeocin or nourseothricin were not resistant against blasticidin-S. In a proof of concept, we also successfully generated double resistant (against blasticidin-S and nourseothricin) cell lines by co-delivering the vector with a vector conferring nourseothricin resistance to wild-type cells.
博来霉素（腐草霉素D1的一种制剂）或诺尔丝菌素。这限制了可以进行的连续遗传转化的数量。为了扩大[模式硅藻名称未给出]的生物技术可能性，我们寻找了可能适用于这种硅藻的其他抗生素和相应的抗性基因。在本研究中测试的三种不同抗生素中，杀稻瘟菌素-S和衣霉素在低浓度下对野生型细胞具有致死性，而伏立康唑对[模式硅藻名称未给出]没有可检测到的影响。测试各自的抗性基因时，我们发现杀稻瘟菌素-S脱氨酶基因（[基因名称未给出]）有效地赋予了[模式硅藻名称未给出]对杀稻瘟菌素-S的抗性。此外，我们可以证明[基因名称未给出]的表达不会导致对博来霉素或诺尔丝菌素的交叉抗性，并且对博来霉素或诺尔丝菌素有抗性的遗传转化细胞系对杀稻瘟菌素-S没有抗性。在概念验证中，我们还通过将[基因名称未给出]载体与赋予野生型细胞诺尔丝菌素抗性的载体共同递送，成功产生了对杀稻瘟菌素-S和诺尔丝菌素双重抗性的[模式硅藻名称未给出]细胞系。
