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Determination of Suitable RT-qPCR Reference Genes for Studies of Gene Functions in Laodelphax striatellus (Fallén)

1
Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences, Key Laboratory of Food Quality and Safety of Jiangsu Province, State Key Laboratory Breeding Base, Nanjing 210014, China
2
National University of Singapore (Suzhou) Research Institute, Department of Biological Sciences, National University of Singapore, Kent Ridge 117543, Singapore
3
College of Chemistry and life Sciences, Zhejiang Normal University, Jinhua 32100, China
*
Authors to whom correspondence should be addressed.
Genes 2019, 10(11), 887; https://doi.org/10.3390/genes10110887
Received: 27 September 2019 / Revised: 25 October 2019 / Accepted: 30 October 2019 / Published: 4 November 2019
The reverse transcription quantitative polymerase chain reaction (RT-qPCR) has been widely used to determine gene functions in Laodelphax striatellus (Fallén) (small brown planthopper). Selection of suitable reference gene(s) for normalizations of RT-qPCR data is critical for reliable results. To date, reports on identification of suitable L. striatellus reference genes are still very limited. L. striatellus is a destructive rice pest and it can transmit multiple viruses, including Rice black-streaked dwarf virus (RBSDV), Rice stripe virus (RSV), and Maize rough dwarf virus (MRDV), to many important cereal crops worldwide. In this study, we examined the stablity of seven selected candidate reference genes in L. striatellus at different developmental stages, in different tissues, in RBSDV- or RSV-infected L. striatellus or in RBSDV-infected and Lssynaptojanin 1 (LsSYNJ1)-silenced L. striatellus. The RT-qPCR data representing individual candidate genes were analyzed using five different methods: the delta Ct method, geNorm, NormFinder, BestKeeper, and the RefFinder algorithm, respectively. The most stable reference gene for the specific condition was selected according to a comprehensive analysis using the RefFinder method. Ribosomal protein L5 (LsRPL5) and LsRPL8 are the most stably expressed genes in L. striatellus at different developmental stages. Alpha-1-tubulin (Lsα-TUB) is the most stably expressed reference gene in different tissues of RBSDV viruliferous (RBSDV-V) or non-viruliferous (RBSDV-NV) L. striatellus. LsRPL8 is the most stably expressed reference gene in RBSDV-V or RSV viruliferous (RSV-V) L. striatellus, while beta-tubulin (Lsβ-TUB) is the most stably expressed reference gene in RBSDV-V and LsSYNJ1-silenced L. striatellus. The selected reference genes were further investigated during analyses of RBSDV P5-1 and P10 gene expression in different tissues from RBSDV-V or RBSDV-NV L. striatellus. The stably expressed reference genes identified in this study will benefit future gene function studies using L. striatellus. View Full-Text
Keywords: Laodelphax striatellus; RT-qPCR; reference gene; rice black-streaked dwarf virus; rice stripe virus Laodelphax striatellus; RT-qPCR; reference gene; rice black-streaked dwarf virus; rice stripe virus
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Wu, W.; Liu, H.; Dong, Y.; Zhang, Y.; Wong, S.-M.; Wang, C.; Zhou, Y.; Xu, Q. Determination of Suitable RT-qPCR Reference Genes for Studies of Gene Functions in Laodelphax striatellus (Fallén). Genes 2019, 10, 887.

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