Selection of Suitable Reference Genes for Gene Expression Normalization Studies in Dendrobium huoshanense

Shanyong Yi; Haibo Lu; Chuanjun Tian; Tao Xu; Cheng Song; Wei Wang; Peipei Wei; Fangli Gu; Dong Liu; Yongping Cai; Bangxing Han

doi:10.3390/genes13081486

Selection of Suitable Reference Genes for Gene Expression Normalization Studies in Dendrobium huoshanense

Genes (Basel). 2022 Aug 19;13(8):1486. doi: 10.3390/genes13081486.

Authors

Shanyong Yi^{1

2}, Haibo Lu², Chuanjun Tian¹, Tao Xu^{1

2}, Cheng Song^{1

2}, Wei Wang^{1

2}, Peipei Wei^{1

2}, Fangli Gu^{1

2}, Dong Liu^{1

2}, Yongping Cai³, Bangxing Han^{1

2}

Affiliations

¹ Department of Biological and Pharmaceutical Engineering, West Anhui University, Lu'an 237012, China.
² Anhui Engineering Laboratory for Conservation and Sustainable Utilization of Traditional Chinese Medicine Resources, West Anhui University, Lu'an 237061, China.
³ College of Life Sciences, Anhui Agricultural University, Hefei 230036, China.

Abstract

Dendrobium huoshanense is a kind of precious herb with important medicinal and edible value in China, which is widely used in traditional Chinese medicine for various diseases. Recent studies have paid close attention to the genetic expression of the biosynthetic pathway of the main active components (polysaccharides, alkaloids, and flavonoids), and real-time polymerase chain reaction (qPCR) is one of the most widely used methods for doing so. However, so far, no reference gene selections have been reported in D. huoshanense. In this study, 15 reference gene candidates (GAPDH, eIF, EF-1α, PP2A, UBCE, RPL5, TBP, APT1, MDH, PTBP3, PEPC, CYP71, NCBP2, TIP41, and F-box) were selected and evaluated for their expression stability in D. huoshanense under various experimental conditions, including in different tissues (root, stem, and leaf), abiotic stresses (oxidative, drought, cold, and UV), and hormone treatment (methyl jasmonate) using three statistical programs (geNorm, NormFinder, and BestKeeper). Then, the RefFinder program was employed to comprehensively validate the stability of the selected reference genes. Finally, the expression profiles of the CESA and GMPP genes were further analyzed, and these results indicated that TBP, NCBP2, and CYP71 were the top three most stable reference genes after comprehensive comparison, which could be used as stable reference genes for normalizing the genes expression in D. huoshanense. This study described here provides the first data regarding on reference gene selection in D. huoshanense, which will be extremely beneficial for future research on the gene expression normalization in D. huoshanense.

Keywords: Dendrobium huoshanense; abiotic stress; qPCR; reference gene; stability evaluation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Dendrobium* / genetics
Droughts
Gene Expression
Genes, Plant*
Stress, Physiological / genetics

Grants and funding

This research was supported by China Agriculture Research System of MOF and MARA, Key Project at Central Government Level: The ability establishment of sustainable use for valuable Chinese medicine resources (2060302), the Natural Science Foundation of Anhui Province (2208085QH266), Postdoctoral Science Foundation of West Anhui University (WXBSH2021002), High level talent project of West Anhui University (WGKQ2021024).