Journal of Integrative Agriculture ›› 2018, Vol. 17 ›› Issue (12): 2745-2757.DOI: 10.1016/S2095-3119(18)61926-4

所属专题： 昆虫和植物互作合辑Insect and Plant Interact

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  • 收稿日期:2017-09-29 出版日期:2018-12-01 发布日期:2018-12-03

Assessment of suitable reference genes for qRT-PCR analysis in Adelphocoris suturalis

LUO Jing1, 2, 3, MA Chao1, LI Zhe1, ZHU Bang-qin3, ZHANG Jiang2, LEI Chao-liang3, JIN Shuang-xia1, J. Joe Hull4, CHEN Li-zhen1, 3   

  1. 1 National Key Laboratory of Crop Genetic Improvement and National Centre of Plant Gene Research, Huazhong Agricultural University, Wuhan 430070, P.R.China
    2 Hubei Collaborative Innovation Center for Green Transformation of Bio-Resources, College of Life Science, Hubei University, Wuhan 430062, P.R.China
    3 Hubei Insect Resources Utilization and Sustainable Pest Management Key Laboratory, College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, P.R.China
    4 U.S. Arid Land Agricultural Research Center, Agricultural Research Service, U.S. Department of Agriculture, Maricopa 85138, USA
  • Received:2017-09-29 Online:2018-12-01 Published:2018-12-03
  • Contact: Correspondence CHEN Li-zhen, E-mail: lzchen@mail.hzau.edu.cn
  • Supported by:
    This work was funded by the National Special Key Project for Transgenic Breeding, China (2016ZX08011002) and the Open Fundation of State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences (SKLOF201415).

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Abstract:

Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is the most commonly-used tool for measurement of gene expression, but its accuracy and reliability depend on appropriate data normalization with the use of one or more stable reference genes.  Adelphocoris suturalis is one of the most destructive pests of cotton, but until recently knowledge of its underlying molecular physiology had been hindered by a lack of molecular resources.  To facilitate research on this pest, we evaluated 12 common housekeeping genes studied in insects (GAPDH, ACT, βACT, TBP, SDH, βTUB, EF1γ, EF1α, EF1δ, RPL32, RPS15, and RPL27) for their expression stability in A. suturalis when subjected to various experimental treatments, including three biotic (developmental stage and sex, tissue type, and metathoracic scent gland for varying developmental stages and sexes) and one abiotic (RNA interference injection) conditions.  Four dedicated algorithms (ΔCt method, geNorm, BestKeeper and NormFinder) were used to analyze gene expression stability.  In addition, RefFinder provided an overall ranking of the stability/suitability of these candidates.  This study is the first to provide a comprehensive list of suitable reference genes for gene expression analyses in A. suturalis, which can serve to facilitate transcript expression study of related biological processes in this and related species.
 

Key words: Adelphocoris suturalis ,  reference gene ,  qRT-PCR ,  normalization ,  expression stability