Selection of stable reference genes for RT–qPCR in Salmo trutta

Sun, Shuaijie; Wang, Zhitong; Yuan, Ding; Ni, Mengke; Xu, Huifen; Wang, Wanliang; Zhang, Chi; Chen, Meiqun; ZhaXi, Lamu; Zhou, Jianshe; Li, Ming

doi:10.1016/j.aqrep.2022.101290

Cited by 1 publication

(4 citation statements)

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“…Furthermore, a similar discrepancy existed between Bestkeeper and NormFinder, Delta Ct, Genorm, and comprehensive analysis when the least unstable reference gene was analyzed. Several studies have mentioned different analytical programs that result in different stability rankings [ 20 , 31 ]. This disparity was caused by differences in the algorithms and procedures used by each computational program.…”

Section: Discussionmentioning

confidence: 99%

“…However, it is yet to be determined whether the expression stability of reference genes has been tested in plant protein-based diet experiments. In the present study, six candidate genes that cover multiple physiological activities of cells involved in cytoskeleton formation ( β-actin ), protein synthesis ( EF1a ), glycolysis canalization ( GAPDH ), purine nucleotide cycle ( HPRT1 ), protein translation ( 18S rRNA ), immune response, and structural protein ( B2M ) were chosen based on previous studies on fish [ 4 , 28 , 31 ]. The expression variations in different tissues reflected by violin maps or the interquartile range of Ct values of each gene indicated the tissue-dependent expression of these reference genes in hybrid yellow catfish fed with different SBM levels.…”

Section: Discussionmentioning

confidence: 99%

“…HPRT1 , a coding gene encoding hypoxanthine-guanine phosphoribosyl transferase (HGPRT) involved in the purine nucleotide cycle, exhibits species- [ 31 , 39 ], tissue-, and test substance-dependent expression [ 20 , 22 ]. Similar findings were found in the current study.…”

Section: Discussionmentioning

confidence: 99%

“…result in different stability rankings [20,31]. This disparity was caused by differences in the algorithms and procedures used by each computational program.…”

Section: Discussionmentioning

confidence: 99%

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Screening and Stability Analysis of Reference Genes for Gene Expression Normalization in Hybrid Yellow Catfish (Pelteobagrus fulvidraco ♀ × Pelteobagrus vachelli ♂) Fed Diets Containing Different Soybean Meal Levels

Guo,

Zhang,

Zhang

et al. 2023

Aquaculture Nutrition

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In this study, we screened the expression stability of six reference genes (18S rRNA, β-actin, GAPDH, EF1a, B2M, and HPRT1) in hybrid yellow catfish (n = 6), considering the SBM levels, sampling time points, and different tissues. Four different statistical programs, BestKeeper, NormFinder, Genorm, and Delta Ct, combined with a method that comprehensively considered all results, were used to evaluate the expression stability of these reference genes systematically. The results showed that SBM levels significantly impacted the expression stability of most of the reference genes studied and that this impact was time-, dose-, and tissue-dependent. The expression stability of these six reference genes varied depending on tissue, sampling time point, and SBM dosage. Additionally, more variations were found among different tissues than among different SBM levels or sampling time points. Due to its high expression, 18S rRNA was excluded from the list of candidate reference genes. β-actin and GAPDH in the liver and β-actin, HPRT1 and EF1a in the intestine were the most stable reference genes when SBM levels were considered. HPRT1, and EF1a in tissues sampled at 2 W and EF1a and β-actin in tissues sampled at 4 and 6 W were proposed as two stable reference genes when different tissues were considered. When the sampling time points were considered, β-actin, EF1a, and HPRT1 were the top three stable reference genes in the intestine. In contrast, β-actin and B2M are the most stable reference genes in the liver. In summary, β-actin, EF1a, and HPRT1 were the more stable reference genes in this study. The stability of reference genes depends on the tissues, sampling time points, and SBM diet levels in hybrid yellow catfish. Therefore, attention should be paid to these factors before selecting suitable reference genes for normalizing the target genes.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…result in different stability rankings [20,31]. This disparity was caused by differences in the algorithms and procedures used by each computational program.…”

Section: Discussionmentioning

confidence: 99%

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