Open AccessReliable reference gene selection for C ordyceps militaris gene expression studies under different developmental stages and media
Author(s) -
Lian Tiantian,
Yang Tao,
Liu Guijun,
Sun Junde,
Dong Caihong
Publication year - 2014
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/1574-6968.12492
Subject(s) - reference genes , cordyceps militaris , biology , gene expression , gene , genetics , gene expression profiling , selection (genetic algorithm) , real time polymerase chain reaction , computational biology , cordyceps , botany , computer science , artificial intelligence
C ordyceps militaris is considered a model organism for the study of C ordyceps species, which are highly prized in traditional C hinese medicine. Gene expression analysis has become more popular and important in studies of this fungus. Reference gene validation under different experimental conditions is crucial for RT ‐q PCR analysis. In this study, eight candidate reference genes, actin , cox5 , gpd , rpb1 , tef1 , try , tub , and ubi , were selected and their expression stability was evaluated in C . militaris samples using four algorithms, genorm , normfinder , bestkeeper, and the comparative ∆ C t method. Three sets of samples, five different developmental stages cultured in wheat medium and pupae, and all the samples pool were included. The results showed that rpb1 was the best reference gene during all developmental stages examined, while the most common reference genes, actin and tub , were not suitable internal controls. Cox5 also performed poorly and was less stable in our analysis. The ranks of ubi and gpd were inconsistent in different sample sets by different methods. Our results provide guidelines for reference gene selection at different developmental stages and also represent a foundation for more accurate and widespread use of RT ‐q PCR in C . militaris gene expression analysis.