Selection of reference genes for quantitative real-time RT-PCR on gene expression in Golden Pompano (Trachinotus ovatus)

Chen, X J; Zhang, X Q; Huang, Shunmou; Cao, Zhenjie; Qin, Qi; Hu, Weiming; Sun, Yuan; Zhou, Yongcan

doi:10.1515/pjvs-2017-0071

Cited by 23 publications

(15 citation statements)

References 27 publications

(28 reference statements)

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“…The ideal reference gene should be stably expressed in all tissues and cell types without being affected by internal or external factors 27 . However, owing to differences in species 45 , 46 , tissue functions 47 , 48 , and metabolic characteristics 49 , 50 , it is almost impossible to obtain universal reference genes.…”

Section: Discussionmentioning

confidence: 99%

“…They are not only involved in protein synthesis, but also in the processes of replication, transcription, RNA processing, DNA repair, self-translational regulation, and developmental regulation. Therefore, these genes can be used as an internal reference genes 47 . The present study showed that the expression levels of RPS29 and RPL19 in different taimen tissues under heat-stress conditions were more stable compared with those of other candidate reference genes.…”

Section: Discussionmentioning

confidence: 99%

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Evaluation of qPCR reference genes for taimen (Hucho taimen) under heat stress

Yang

Tong

Dong

et al. 2022

Sci Rep

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As a powerful and attractive method for detecting gene expression, qRT-PCR has been broadly used in aquaculture research. Understanding the biology of taimen (Hucho taimen) has drawn increasing interest because of its ecological and economic value. Stable reference genes are required for the reliable quantification of gene expression, but such genes have not yet been optimized for taimen. In this study, the stability levels of 10 commonly used candidate reference genes were evaluated using geNorm, NormFinder, BestKeeper, and RefFinder. The expression levels of the 10 genes were detected using 240 samples from 48 experimental groups consisting of 40 individuals treated under four heat-stress conditions (18, 20, 22, and 24 °C) for 24 h and 26 °C for 4, 24, 48, and 72 h. Six tissues (blood, heart, brain, gill, skin, and liver) were collected from each individual. Ribosomal protein S29 (RPS29) and ribosomal protein L19 (RPL19) were the most stable genes among all of the samples, whereas 28S ribosomal RNA (28S rRNA), attachment region binding protein (ARBP), and 18S ribosomal RNA (18S rRNA) were the least stable. These results were verified by an expression analysis of taimen heat-stress genes (heat shock protein 60, hsp60, and heat shock protein 70, hsp70). In conclusion, RPS29 and RPL19 are the optimal reference genes for qRT-PCR analyses of taimen, irrespective of the tissue and experimental conditions. These results allow the reliable study of gene expression in taimen.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Evaluation of qPCR reference genes for taimen (Hucho taimen) under heat stress

Yang

Tong

Dong

et al. 2022

Sci Rep

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“…GAPDH is sensitive to changes in cellular homeostasis, and its expression is modified at tissue level during developmental and physiological conditions (Infante et al, 2008). Unsuitability of GAPDH as RG has been reported in numerous experiments: larval development in Senegalese sole and Atlantic halibut (Infante et al, 2008), bacterial infection in Japanese flounder (Zheng & Sun, 2011), salinity stress in spotted seabass (Wang et al, 2018), developmental stages in zebra fish (McCurley & Callard, 2008) and loach (Chen et al, 2017) and gonadal development in rice eel (Hu et al, 2014). Conversely, some studies have found GAPDH as a stable RG in different embryonic developmental stages of half‐smooth tongue sole (Liu et al, 2014) and magur (Mir et al, 2018), and during myxosporean infection in rainbow trout (Kosakyan et al, 2019).…”

Section: Discussionmentioning

confidence: 99%

Identification of reference genes for quantitative expression analysis in Indian catfish,Clarias magur, under physiological and pathological conditions

Muduli

Rathore

Singh

et al. 2022

Aquaculture Research

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The reverse transcription quantitative real‐time PCR (RT‐qPCR) is widely preferred tool for expression analysis of target gene at mRNA level. However, in order to obtain significant biological comparison, data normalization to a stable endogenous internal control gene also called reference gene (RG) is of utmost importance. In the present study, we evaluated the expression stability of four commonly used housekeeping genes, that is beta actin, β‐actin; glyceraldehyde‐3‐phosphate dehydrogenase, GAPDH; elongation factor‐1 alpha, EF‐1α and 18S ribosomal RNA, 18S for qPCR data normalization. This study was conducted for two varying physiological (Group I: fed vs. non‐fed and Group II: earthen pond vs. fibre reinforced plastic tank reared) and one pathological condition (Group III: Aeromonas hydrophila injected vs. PBS injected) in seven different tissues of Indian catfish, Clarias magur, an economically valuable food fish of South‐East Asia. Tissue wise expression stability analysis using web‐based comprehensive tool Reffinder revealed that EF‐1α was the most stable RG in kidney, liver, spleen and gills of group I; spleen, intestine, muscle and brain in group II; and kidney, liver, intestine, muscle and brain in group III. β‐actin was the most stable RG in intestine of group I; liver and brain in group II; spleen and gill in group III. 18S was the most stable RG in muscle and brain of group I; kidney and gill in group II. GAPDH was the least stable RG among four selected genes under all the experimental conditions. These findings on selection of RGs will lead to precise interpretation of relative gene expression in different tissues by RT‐qPCR in C. magur in the context of physiological studies and bacterial infection.

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“…The ideal reference gene should be stably expressed in all the tissues and cell types without being affected by internal or external factors 13 . However, owing to differences in species 25,26 , tissue functions 27,28 and metabolic characteristics 29,30 , it is almost impossible to obtain universal reference genes. Many researchers optimise the reference genes used for different species.…”

Section: Discussionmentioning

confidence: 99%

“…RPS29, RPL13 and RPL19 encode large ribosomal subunits, and they are highly conserved in eukaryotes. They are not only involved in protein synthesis, but also in the processes of replication, transcription, RNA processing, DNA repair, selftranslational regulation and developmental regulation; therefore, they can be used as an internal reference genes 27 . The present study showed that the expression levels of RPS29 and RPL19 in different taimen tissues under heat-stress conditions were more stable compared with those of other candidate reference genes.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of qPCR Reference Genes For Taimen (Hucho Taimen)

Yang

Tong

Dong

et al. 2021

Preprint

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As a powerful and attractive method for detecting gene expression, qRT-PCR has been broadly used in aquacultural research. Understanding the biology of taimen (Hucho taimen) has attracted increased interest because of its ecological and economic values. Stable reference genes are required for the reliable quantification of gene expression, but such genes have not yet been optimised for taimen. In this study, the stability levels of 10 commonly used candidate reference genes were evaluated using four methods, geNorm, NormFinder, BestKeeper and RefFinder. The expression levels of the 10 genes were detected using samples from 48 experimental groups that consisted of six tissues (blood, heart, brain, gill, skin and liver) collected under five heat-stress conditions (18, 20, 22, 24 and 26°C) and four heat-stress processing times (4, 24, 48 and 96 h). RPS29 and RPL19 were the most stable genes among all the samples, whereas 28S rRNA, ARBPR and 18S rRNA were the least stable. These results were verified by an expression analysis of the heat-stress genes (hsp60 and hsp70) of taimen. In conclusion, RPS29 and RPL19 are the optimal reference genes for qRT-PCR analyses of taimen, irrespective of the tissue and experimental conditions, and these results allow the reliable study of gene expression in taimen.

show abstract

Selection of reference genes for quantitative real-time RT-PCR on gene expression in Golden Pompano (Trachinotus ovatus)

Cited by 23 publications

References 27 publications

Evaluation of qPCR reference genes for taimen (Hucho taimen) under heat stress

Evaluation of qPCR reference genes for taimen (Hucho taimen) under heat stress

Identification of reference genes for quantitative expression analysis in Indian catfish,Clarias magur, under physiological and pathological conditions

Evaluation of qPCR Reference Genes For Taimen (Hucho Taimen)

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