Selection and validation of reference genes for quantitative gene expression normalization in Taxus spp.
Kaikai Zhang1,2, Wei Fan3, Duanfen Chen2
1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of State Forestry Administration, The Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091, China.
Scientific Reports
|December 18, 2020
Summary
This study identifies stable reference genes, GAPDH1 and SAND, for accurate gene expression analysis in Taxus species using quantitative real-time PCR (qRT-PCR). These findings are crucial for understanding taxol biosynthesis and improving experimental reproducibility in related species.
Area of Science:
- Plant Molecular Biology
- Biotechnology
- Pharmacognosy
Background:
- Quantitative real-time PCR (qRT-PCR) is essential for gene expression studies, requiring stable reference genes for accurate quantification.
- Taxus species are vital sources of the anticancer compound taxol, making their gene expression analysis critical for understanding biosynthesis.
- Limited research exists on suitable reference genes for qRT-PCR in Taxus species.
Purpose of the Study:
- To screen and validate reference genes for reliable gene expression normalization in Taxus species.
- To establish robust qRT-PCR protocols for studying taxol biosynthesis and related genes.
- To provide essential tools for accurate gene expression analysis in Taxus species under various conditions.
Main Methods:
- Selection of eight candidate reference genes for qRT-PCR analysis.
- Application of statistical algorithms (geNorm, NormFinder, BestKeeper, ΔCt, RefFinder) for gene stability assessment.
- Analysis of gene expression data from Taxus × media cell lines and Taxus chinensis var. mairei tissues under diverse experimental conditions.
Main Results:
- Identification of GAPDH1 and SAND as the most stable reference genes across different treatments and tissues in Taxus species.
- Demonstration of significant differences in TcMYC expression under salicylic acid treatment, highlighting the need for validated reference genes.
- Validation of the selected reference genes for accurate normalization of qRT-PCR data in Taxus species.
Conclusions:
- GAPDH1 and SAND are recommended as reliable reference genes for qRT-PCR in Taxus species, ensuring data accuracy and reproducibility.
- This study provides a foundation for future research into taxol biosynthesis and gene regulation in Taxus.
- The findings are applicable to other related plant species, advancing molecular studies in ethnopharmacology and plant biotechnology.
