Genome-Wide Screening and Stability Verification of the Robust Internal Control Genes for RT-qPCR in Filamentous Fungi
文献类型： 外文期刊
作者： Yang, Yayong 1 ; Xu, Xinyu 1 ; Jing, Zhuohan 1 ; Ye, Jun 1 ; Li, Hui 3 ; Li, Xiaoyu 1 ; Shi, Lei 1 ; Chen, Mengyu 1 ; Wang, Tengyun 1 ; Xie, Baogui 2 ; Tao, Yongxin 1 ;
作者机构： 1.Fujian Agr & Forestry Univ, Coll Hort, Fuzhou 350002, Peoples R China
2.Fujian Agr & Forestry Univ, Coll Life Sci, Mycol Res Ctr, Fuzhou 350002, Peoples R China
3.Hebei Acad Agr & Forestry Sci, Inst Cash Crops, Shijiazhuang 050051, Hebei, Peoples R China
关键词： internal control genes; filamentous fungi; RT-qPCR; gene expression
期刊名称：JOURNAL OF FUNGI （ 影响因子：5.724； 五年影响因子：6.413 ）
ISSN：
年卷期： 2022 年 8 卷 9 期
页码：
收录情况： SCI
摘要： In real-time quantitative PCR (RT-qPCR), internal control genes (ICGs) are crucial for normalization. This study screened 6 novel ICGs: Pre-mRNA-splicing factor cwc15 (Cwf15); ER associated DnaJ chaperone (DnaJ); E3 ubiquitin-protein ligase NEDD4 (HUL4); ATP-binding cassette, subfamily B (MDR/TAP), member 1 (VAMP); Exosome complex exonuclease DIS3/RRP44 (RNB); V-type H+-transporting ATPase sub-unit A (V-ATP) from the 22-transcriptome data of 8 filamentous fungi. The six novel ICGs are all involved in the basic biological process of cells and share the different transcription levels from high to low. In order to further verify the stability of ICGs candidates, the six novel ICGs as well as three traditional housekeeping genes: beta-actin (ACTB); beta-tubulin (beta-TUB); glyceraldehyde-3-phosphate dehydrogenase gene (GAPDH) and the previously screened reference genes: SPRY-domain-containing protein (SPRYp); Ras-2 protein (Ras); Vacuolar protein sorting protein 26 (Vps26) were evaluated by geNorm and NormFinder statistical algorithms. RT-qPCR of 12 ICGs were performed at different developmental stages in Flammulina filiformis and under different treatment conditions in Neurospora crassa. The consistent results of the two algorithms suggested that the novel genes, RNB, V-ATP, and VAMP, showed the highest stability in F. filiformis and N. crassa. RNB, V-ATP, and VAMP have high expression stability and universal applicability and therefore have great potential as ICGs for standardized calculation in filamentous fungi. The results also provide a novel guidance for the screening stable reference genes in RT-qPCR and a wide application in gene expression analysis of filamentous fungi.
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