Assessment of reference gene stability in Rice stripe virus and Rice black streaked dwarf virus infection rice by quantitative Real-time PCR.
BACKGROUND
Stably expressed reference gene(s) normalization is important for the understanding of gene expression patterns by quantitative Real-time PCR (RT-qPCR), particularly for Rice stripe virus (RSV) and Rice black streaked dwarf virus (RBSDV) that caused seriously damage on rice plants in China and Southeast Asia.


METHODS
The expression of fourteen common used reference genes of Oryza sativa L. were evaluated by RT-qPCR in RSV and RBSDV infected rice plants. Suitable normalization reference gene(s) were identified by geNorm and NormFinder algorithms.


RESULTS
UBQ 10 + GAPDH and UBC + Actin1 were identified as suitable reference genes for RT-qPCR normalization under RSV and RBSDV infection, respectively. When using multiple reference genes, the expression patterns of OsPRIb and OsWRKY, two virus resistance genes, were approximately similar with that reported previously. Comparatively, by using single reference gene (TIP41-Like), a weaker inducible response was observed.


CONCLUSIONS
We proposed that the combination of two reference genes could obtain more accurate and reliable normalization of RT-qPCR results in RSV- and RBSDV-infected plants. This work therefore sheds light on establishing a standardized RT-qPCR procedure in RSV- and RBSDV-infected rice plants, and might serve as an important point for discovering complex regulatory networks and identifying genes relevant to biological processes or implicated in virus.
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China. 2011116138@njau.edu.cn.|Scientific Observing and Experimental Station of Crop Pests in Nanjing, Ministry of Agriculture, China, Nanjing, 210014, China. 2011116138@njau.edu.cn.|College of Life Sciences, Nanjing Agricultural University, Nanjing, 210095, China. 2011116138@njau.edu.cn.
Peng
P
Fang
College of Life Sciences, Nanjing Agricultural University, Nanjing, 210095, China. 2014116011@njau.edu.cn.
Rongfei
R
Lu
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China. sunfeng1201@gmail.com.|Scientific Observing and Experimental Station of Crop Pests in Nanjing, Ministry of Agriculture, China, Nanjing, 210014, China. sunfeng1201@gmail.com.
Feng
F
Sun
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China. lanying0918@hotmail.com.|Scientific Observing and Experimental Station of Crop Pests in Nanjing, Ministry of Agriculture, China, Nanjing, 210014, China. lanying0918@hotmail.com.
Ying
Y
Lan
College of Life Sciences, Nanjing Agricultural University, Nanjing, 210095, China. wbshenh@njau.edu.cn.
Wenbiao
W
Shen
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China. dulinlin12@126.com.|Scientific Observing and Experimental Station of Crop Pests in Nanjing, Ministry of Agriculture, China, Nanjing, 210014, China. dulinlin12@126.com.
Linlin
L
Du
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China. yjzhou@jaas.ac.cn.|Scientific Observing and Experimental Station of Crop Pests in Nanjing, Ministry of Agriculture, China, Nanjing, 210014, China. yjzhou@jaas.ac.cn.
Yijun
Y
Zhou
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Nanjing, 210014, China. zhoutong@jaas.ac.cn.|Scientific Observing and Experimental Station of Crop Pests in Nanjing, Ministry of Agriculture, China, Nanjing, 210014, China. zhoutong@jaas.ac.cn.
Tong
T
Zhou
D002681:China; D020869:Gene Expression Profiling / Q000592:standards*; D042822:Genomic Instability; D012275:Oryza / Q000821:virology*; D010935:Plant Diseases / Q000821:virology*; D060888:Real-Time Polymerase Chain Reaction / Q000592:standards*; D012015:Reference Standards; D012087:Reoviridae / Q000235:genetics / Q000302:isolation & purification*; D029364:Tenuivirus / Q000235:genetics / Q000302:isolation & purification*