The optimal reference gene validation in Cyclocarya paliurus (Batal.) Iljinskaja under environmental stresses

Abstract

Cyclocarya paliurus (Batal.) Iljinskaja is a special plant species grown at high elevation in southwestern China. The bark, leaves, and roots are used for many purposes including reduction of fever and enhancement of blood insulin level. However, its growth is usually limited by the fragile environment and climate variability. Previous studies attributed the environment‐relevant yield reductions to stress induced gene expression changes. The accuracy and reliability of reverse transcription quantitative polymerase chain reaction (RT‐qPCR), a commonly accepted gene expression method, depends on the choice of appropriate reference genes, which are used for gene expression normalization. However, there are no reports at present about reference genes of C. paliurus under environmental stresses. Therefore, this study was conducted to identify reference genes for C. paliurus under environmental stresses. In this work, we selected seven common reference genes as our candidates: ACT2, ACTF, GAPDH, TUA, TUB, UBQ4, and 18SrRNA. The stability of these seven reference genes in C. paliurus was quantified by RT‐qPCR and evaluated by geNorm and NormFinder algorithms. The results suggest that although there were variations of the stability of the internal reference genes, two of them were usually good enough for the normalization purpose for low temperature (4 °C), high salinity (NaCl), and drought stress (polyethylene glycol [PEG] treatment). However, these genes should not be used for high‐temperature (42 °C) stress. This study provides insight for reference gene selection under experimental stresses in C. paliurus. It also sets the foundation of wide and accurate use of RT‐qPCR under environmental stresses.

Agricultural sciences and the environment: Reviewing recent technologies and innovations to combat the challenges of climate change, environmental protection, and food security

“…(2022) and in Cyclocarya paliurus Iljinskaja, a special plant species grown at high elevation in southwestern China, by Yang, Fan, et al. (2022). The authors noted that GAPDH gene was identified as the best reference gene.…”

Identification of CpbZIP11 in Cyclocarya paliurus Involved in Environmental Stress Responses

“…The cDNA synthesis was carried out using Moloney murine leukemia virus (M-MLV) reverse transcriptase from Invitrogen, following the instructions provided by the manufacturer. To ensure precise quantification, the 18S RNA was utilized as a normalization control for the quantity of cDNA [40]. RT-PCR analysis was performed to investigate the splicing of CpbZIP11.…”

Genome-wide identification and analysis of bZIP gene family reveal their roles during development and drought stress in Wheel Wingnut (Cyclocarya paliurus)

“…Furthermore, there is evidence for strong health-promoting effects of its leaves, including the ability to lower blood sugar, reduce blood lipids, protect against cancer, and enhance immunity [ 24 ]. The growth and development of C. paliurus leaves are affected by environmental stress, such as drought, salt, cold, and heat [ 25 ], and various TFs contribute to the regulation of growth in C. paliurus leaves. For example, bZIP is involved in the regulation of amino acid biosynthesis [ 26 ], and MYB and bHLH are involved in the regulation of flavonoid biosynthesis [ 27 ].…”