Development of an Expression Vector to Overexpress or Downregulate Genes in Curvularia protuberata.
Development of an Expression Vector to Overexpress or Downregulate Genes in Curvularia protuberata.
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DOI：
10.3390/jof4020054
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发表时间：
2018-05-05
期刊：
影响因子：
--
通讯作者：
Morsy M
中科院分区：
文献类型：
--
作者：
Liu C;Cleckler B;Morsy M
凸弯孢菌是子囊菌门的一种内生真菌，只有当它携带一种真菌病毒（称为弯孢耐热病毒（CThTV））时，才能为植物提供耐热性，并在这些生物体之间形成三方共生关系。在热胁迫下，几个基因的表达差异之间的脱毒C。rubbulata（VF）和C. prolongata携带CThTV（AN）。我们构建了一个表达载体pM2 Z-fun，该载体携带由ToxA启动子驱动的zeocin抗性基因，用于研究C.为了更好地理解这种三方共生。利用这个新的3.7kb的载体，在C.包括参与海藻糖、黑色素和过氧化氢酶生物合成途径的基因在VF或ANC中成功地过表达或下调。prolongata菌株。VF过表达株的代谢产物和酶活性高于对照VF株。此外，在AN菌株中相同基因的表达下调导致比对照AN菌株中更低的代谢产物和酶活性。新构建的表达载体pM2 Z-fun已成功地用于表达目的基因。在其他子囊菌门真菌中的进一步功能表达研究中将是有用的。
Curvularia protuberata, an endophytic fungus in the Ascomycota, provides plants with thermotolerance only when it carries a mycovirus known as Curvularia thermotolerance virus (CThTV), and forms a three-way symbiotic relationship among these organisms. Under heat stress, several genes are expressed differently between virus-free C. protuberata (VF) and C. protuberata carrying CThTV (AN). We developed an expression vector, pM2Z-fun, carrying a zeocin resistance gene driven by the ToxA promoter, to study gene functions in C. protuberata to better understand this three-way symbiosis. Using this new 3.7-kb vector, five genes that are differentially expressed in C. protuberata—including genes involved in the trehalose, melanin, and catalase biosynthesis pathways—were successfully overexpressed or downregulated in VF or AN C. protuberata strains, respectively. The VF overexpression lines showed higher metabolite and enzyme activity than in the control VF strain. Furthermore, downregulation of expression of the same genes in the AN strain resulted in lower metabolite and enzyme activity than in the control AN strain. The newly generated expression vector, pM2Z-fun, has been successfully used to express target genes in C. protuberata and will be useful in further functional expression studies in other Ascomycota fungi.
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