Reference Genes Screening and Gene Expression Patterns Analysis Involved in Gelsenicine Biosynthesis under Different Hormone Treatments in Gelsemium elegans

Abstract

Reverse transcription quantitative polymerase chain reaction (RT-qPCR) is an accurate method for quantifying gene expression levels. Choosing appropriate reference genes to normalize the data is essential for reducing errors. Gelsemium elegans is a highly poisonous but important medicinal plant used for analgesic and anti-swelling purposes. Gelsenicine is one of the vital active ingredients, and its biosynthesis pathway remains to be determined. In this study, G. elegans leaf tissue with and without the application of one of four hormones (SA, MeJA, ETH, and ABA) known to affect gelsenicine synthesis, was analyzed using ten candidate reference genes. The gene stability was evaluated using GeNorm, NormFinder, BestKeeper, ∆CT, and RefFinder. The results showed that the optimal stable reference genes varied among the different treatments and that at least two reference genes were required for accurate quantification. The expression patterns of 15 genes related to the gelsenicine upstream biosynthesis pathway was determined by RT-qPCR using the relevant reference genes identified. Three genes 8-HGO, LAMT, and STR, were found to have a strong correlation with the amount of gelsenicine measured in the different samples. This research is the first study to examine the reference genes of G. elegans under different hormone treatments and will be useful for future molecular analyses of this medically important plant species.

Selection and Verification of Standardized Reference Genes of Angelica dahurica under Various Abiotic Stresses by Real-Time Quantitative PCR

“…Moreover, previous studies have indicated that plants subjected to abiotic stress exhibit increased yields of secondary metabolites, accompanied by changes in the expression levels of relevant biosynthetic genes [23][24][25]. It has been demonstrated that the application of methyl jasmonate (MeJA) leads to an augmentation in the synthesis and accumulation of tanshinones and phenolic acids within the hairy roots of Salvia miltiorrhiza.…”

Evaluation of Reference Genes for Normalizing RT-qPCR and Analysis of the Expression Patterns of WRKY1 Transcription Factor and Rhynchophylline Biosynthesis-Related Genes in Uncaria rhynchophylla

“…In Euodia rutaecarpa, the expression patterns of HMGCR, SQE, and CYP450 calculated by screened reference genes were consistent with the content of limonin, thus implying their potential involvement in the limonin biosynthesis of E. rutaecarpa. Furthermore, qRT-PCR was used to analyze the expression patterns of 15 genes involved in the terpenoid indole alkaloids pathway using stable reference genes, and three key enzyme genes were found to have strong correlations with the gelsenicine in Gelsemium elegans [13,14]. qRT-PCR has high sensitivity, high specificity, and simple operation [15].…”