Evaluation and validation of suitable reference genes for quantitative real-time PCR analysis in lotus (Nelumbo nucifera Gaertn.).

Publisher: Nature Publishing Group Country of Publication: England NLM ID: 101563288 Publication Model: Electronic Cited Medium: Internet ISSN: 2045-2322 (Electronic) Linking ISSN: 20452322 NLM ISO Abbreviation: Sci Rep Subsets: MEDLINE

Original Publication: London : Nature Publishing Group, copyright 2011-

Real-Time Polymerase Chain Reaction*/standards ; Real-Time Polymerase Chain Reaction*/methods ; Nelumbo*/genetics ; Genes, Plant* ; Gene Expression Regulation, Plant*; Reference Standards ; Gene Expression Profiling/methods ; Gene Expression Profiling/standards ; Lotus/genetics ; Lotus/growth & development

The qRT-PCR technique has been regarded as an important tool for assessing gene expression diversity. Selection of appropriate reference genes is essential for validating deviation and obtaining reliable and accurate results. Lotus (Nelumbo nucifera Gaertn) is a common aquatic plant with important aesthetic, commercial, and cultural values. Twelve candidate genes, which are typically used as reference genes for qRT-PCR in other plants, were selected for this study. These candidate reference genes were cloned with, specific primers designed based on published sequences. In particular, the expression level of each gene was examined in different tissues and growth stages of Lotus. Notably, the expression stability of these candidate genes was assessed using the software programs geNorm and NormFinder. As a result, the most efficient reference genes for rootstock expansion were TBP and UBQ. In addition, TBP and EF-1α were the most efficient reference genes in various floral tissues, while ACT and GAPDH were the most stable genes at all developmental stages of the seed. CYP and GAPDH were the best reference genes at different stages of leaf development, but TUA was the least stable. Meanwhile, the gene expression profile of NnEXPA was analyzed to confirm the validity of the findings. It was concluded that, TBP and GAPDH were identified as the best reference genes. The results of this study may help researchers to select appropriate reference genes and thus obtain credible results for further quantitative RT-qPCR gene expression analyses in Lotus.
(© 2024. The Author(s).)

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No.2019ABA108,No.2021BBA093 Technology Innovation Project of Hubei Province of China (Major Program); No.2019ABA108,No.2021BBA093 Technology Innovation Project of Hubei Province of China (Major Program); No. 2020ED73 National modern Agricultural Industrial Technology System (China Ministry of Finance and Ministry of Agriculture and Rural Affairs) and  Talent project of Fuzhou, Jiangxi; No. 2020ED73 National modern Agricultural Industrial Technology System (China Ministry of Finance and Ministry of Agriculture and Rural Affairs) and  Talent project of Fuzhou, Jiangxi

Keywords: Gene expression; Lotus; NormFinder; Reference gene; geNorm; qRT-PCR

Date Created: 20240513 Date Completed: 20240513 Latest Revision: 20240516

20250114

PMC11091181

10.1038/s41598-024-61806-9

38740848