Selection and Validation of Appropriate Reference Genes for Real-Time Quantitative PCR Analysis in Needles of Larix olgensis under Abiotic Stresses

Abstract

Larix olgensis Henry is an important afforestation species in northeastern China because of its fast juvenile growth, high-quality timber, and significant economic and ecological values. The selection of appropriate reference genes is necessary for the normalization of gene expression determination during quantitative real-time polymerase chain reaction (qRT-PCR) experiments. In this study, qRT-PCR was used to study gene expression. Three software packages geNorm, NormFinder, BestKeeper were used, and a comprehensive ranking of candidate reference genes was produced based on their output to evaluate the expression stability of 16 candidate reference genes from L. olgensis under drought, salt, cold, and heat stress. PP2A-1 and GAPDH ranked as the most stable reference genes under drought and cold stress, PP2A-1 and UBQ10 were most stable under salt stress, and TIP41 and ACT2 were most stable under heat stress. The least stable gene was ADP, which ranked the last under all treatments. Expression profile analysis of the antioxidant gene CAT using the two most stable and the single least stable reference genes under each stress further verified that the selected reference genes were suitable for gene expression normalization. This study provides an important foundation for the selection of suitable reference genes for the normalization and quantification of L. olgensis gene expression under abiotic stress conditions.

Effects of stand age and soil organic matter quality on soil bacterial and fungal community composition in Larix gmelinii plantations, Northeast China

“…In Northeast China, afforestation accounts for 70% of the total forest area (Yang et al, 2010). Larix(L.) gmelinii has been widely used as a plantation species because of its rapid growth and excellent wood quality (Li et al, 2020; Yu & Liu, 2020). It is crucial to understand the changes of microbial communities in returning farmland to forest for the ecological functions of this ecosystems.…”

Selection of reference genes for RT-qPCR analysis of rice with Rhizoctonia solani infection and PGPR/KSi application

“…Similar to geNorm, a lower stability value in NormFinder indicates more stable RG. NormFinder calculates not only overall expression variations but also sample subsets, and the relative expression is calculated from the 2 − DCt value [32]. BestKeeper program uses standard deviation, coe cient of variance and correlation coe cient of Cq values of RGs [24].…”