Department of Molecular Biology, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynská dolina, Ilkovičova 6, 842 15, Bratislava 4, Slovak Republic.
J Appl Genet. 2020 May;61(2):275-286. doi: 10.1007/s13353-020-00544-w. Epub 2020 Jan 21.
Yeast Candida utilis is considered to be a potentially advantageous expression system for production of recombinant proteins utilizable for industrial and pharmaceutical purposes. As the scientific literature is not consistent in the ploidy of this yeast, in this work, we focused on resolving the problem via several methods such as the copy number determination of maltase gene by multiplex PCR, measuring α-glucosidase activity, the characterization of maltase gene copy number in deletion mutants using qPCR and flow cytometry. In context with the published data and results obtained in this study about the copy number of the maltase gene on C. utilis genome, we attempted to hypothesise and made conclusion about the ploidy of C. utilis. The results of this work, besides the biotechnological aspect, contribute to the elementary knowledge of C. utilis. The exact information about the ploidy or more specifically about the copy number of appropriate gene is essential for expression cassette dosage determination integrated into the chromosome of the host. In this study, we come to the conclusion that the maltase gene is present in C. utilis genome in four alleles, and in combination with flow cytometry, published information and the published genome sequences, the observations support the theory about tetraploidy of C. utilis.
酵母 Candida utilis 被认为是一种具有潜在优势的表达系统，可用于生产可用于工业和制药目的的重组蛋白。由于该酵母的倍性在科学文献中并不一致，因此在这项工作中，我们通过多重 PCR 测定麦芽糖酶基因的拷贝数、测量α-葡萄糖苷酶活性、使用 qPCR 和流式细胞术表征缺失突变体中的麦芽糖酶基因拷贝数等几种方法来解决这个问题。根据关于 C. utilis 基因组中麦芽糖酶基因拷贝数的已发表数据和本研究的结果，我们尝试对 C. utilis 的倍性进行假设和得出结论。除了生物技术方面，这项工作的结果还为 C. utilis 的基础知识做出了贡献。有关合适基因的倍性或更具体地说有关其拷贝数的准确信息对于整合到宿主染色体中的表达盒剂量确定至关重要。在这项研究中，我们得出结论，麦芽糖酶基因存在于 C. utilis 基因组的四个等位基因中，结合流式细胞术、已发表的信息和已发表的基因组序列，这些观察结果支持关于 C. utilis 四倍体的理论。
