Selection and Validation of Reference Genes in Dendrocalamus brandisii for Quantitative Real-Time PCR

Dendrocalamus brandisii (Munro) Kurz is a sympodial bamboo species with a wide distribution in tropical and subtropical regions. Due to its remarkable regenerative ability and exceptional flavor, this species plays a pivotal role in bolstering the economies of numerous nations across these regions. We recently published a high-quality genome of this species. To date, no study results have identified the optimal reference genes for quantitative real-time polymerase chain reaction (qRT-PCR) normalization in Dendrocalamus brandisii. qRT-PCR offers a highly accurate and effective approach to analyzing gene expression. However, the precision of the resulting quantitative data hinges on the correct choice of reference genes. Twenty-one potential reference genes were identified from the D. brandisii transcriptomes. Their expression in 23 samples, including 8 different tissue organs and 15 samples of D. brandisii under various treatment conditions, were evaluated through qRT-PCR. Subsequently, four software programs-Delta CT, geNorm, NormFinder, and RefFinder-were employed to compare their expression stability. The results revealed that the selection of optimal reference genes varied based on the particular organ and condition being examined. EF-1-α-2 consistently exhibits the most stable expression across diverse tissues, while ACTIN-1, TUBULIN-1, and EF-1-α-2 were the most consistent reference genes in roots, culms, and leaves under various treatments, respectively. In this study, we identified and characterized appropriate internal genes utilized for calibrating qRT-PCR analyses of D. brandisii across different tissue organs and under various treatments. This research will provide key insights for advancing the study of gene functionality and molecular biology in D. brandisii and related species.