Reference Gene Selection for RT-qPCR Analysis on <i>Cucumis melo</i>

ZHANG Lizhen; HAN Xiaoyun; WU Jinghua; Gefu WANG-PRUSKI; ZHANG Zhizhong

doi:10.19303/j.issn.1008-0384.2020.11.002

Volume 35 Issue 11

Nov. 2020

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Article Navigation > Fujian Journal of Agricultural Sciences > 2020 > 35(11): 1179-1187

ZHANG L Z, HAN X Y, WU J H, et al. Reference Gene Selection for RT-qPCR Analysis on Cucumis melo [J]. Fujian Journal of Agricultural Sciences，2020，35（11）：1179−1187 doi: 10.19303/j.issn.1008-0384.2020.11.002

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ZHANG L Z, HAN X Y, WU J H, et al. Reference Gene Selection for RT-qPCR Analysis on Cucumis melo [J]. Fujian Journal of Agricultural Sciences，2020，35（11）：1179−1187 doi: 10.19303/j.issn.1008-0384.2020.11.002

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Reference Gene Selection for RT-qPCR Analysis on Cucumis melo

doi: 10.19303/j.issn.1008-0384.2020.11.002

1.
Joint FAFU-Dalhousie Lab, College of Horticulture, Fujian Agriculture and Forestry University, Fuzhou, Fujian　350002, China
2.
Faculty of Agriculture, Dalhousie University, Truro, NS, Canada　B2N5E3

Received Date: 2020-07-29
Rev Recd Date: 2020-10-04

Available Online: 2020-11-13

Publish Date: 2020-11-30

Abstract

Abstract

Objective In search for internal reference genes of Cucumis melo L. that could stably express in different tissues and under stress conditions to warrant accuracy and reliability of the RT-qPCR analysis on target gene expression. Method Expression stabilities of 9 candidate genes, 18srRNA, TUA, EF1a, Actin1, Actin2, Actin3, Actin4, CYC, and UBI-ep, from the roots, leaves, seeds, and fruits of Xinyinhui melon being treated by water, cinnamic acid, saline alkali or ABA were determined by RT-qPCR and analyzed using the BestKeeper, NormFinder, and geNorm software. Result In different tissues, the top 5 choice genes identified by BestKeeper ranked as CYC＞18s rRNA＞UBI-ep＞EF1a＞TUA, those by NormFinder EF1a＞UBIep＞Actin4＞CYC＞Actin3, and those by geNorm Actin4=Actin3＞Actin1＞EF1a＞UBI-ep. Under various stresses, they were 18s rRNA＞Actin3＞Actin4＞EF1a＞UBI-ep as ranked by BestKeeper, EF1a＞UBI-ep＞Actin4＞CYC＞18s rRNA by NormFinder, and EF1a=UBI-ep＞Actin4＞CYC＞Actin by geNorm. Overall, EF1a appeared to be most stable among the 9 genes. Insofar as variety of tissues is concerned, Actin4, Actin3, Actin1, and EF1a were more stable than the others; and, under stress, EF1a and UBI-ep tended to be superior. Conclusion Stably expressed in the tissues under the stresses as tested, EF1a was selected as the reference gene for studies on C. melo to reduce experimental errors. To further ensure accuracy, application of dual reference genes in RT-qPCR analysis might be considered.
- Cucumis melo L.,
- reference gene,
- RT-qPCR,
- Tissuestissues and organs,
- stress

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