Identifying suitable reference genes for gene expression analysis in developing skeletal muscle in pigs

The selection of suitable reference genes is crucial to accurately evaluate and normalize the relative expression level of target genes for gene function analysis. However, commonly used reference genes have variable expression levels in developing skeletal muscle. There are few reports that systematically evaluate the expression stability of reference genes across prenatal and postnatal developing skeletal muscle in mammals. Here, we used quantitative PCR to examine the expression levels of 15 candidate reference genes (ACTB,GAPDH,RNF7,RHOA,RPS18,RPL32,PPIA,H3F3,API5,B2M,AP1S1,DRAP1,TBP,WSB, andVAPB) in porcine skeletal muscle at 26 different developmental stages (15 prenatal and 11 postnatal periods). We evaluated gene expression stability using the computer algorithms geNorm, NormFinder, and BestKeeper. Our results indicated thatGAPDHandACTBhad the greatest variability among the candidate genes across prenatal and postnatal stages of skeletal muscle development.RPS18,API5, andVAPBhad stable expression levels in prenatal stages, whereasAPI5,RPS18,RPL32, andH3F3had stable expression levels in postnatal stages.API5andH3F3expression levels had the greatest stability in all tested prenatal and postnatal stages, and were the most appropriate reference genes for gene expression normalization in developing skeletal muscle. Our data provide valuable information for gene expression analysis during different stages of skeletal muscle development in mammals. This information can provide a valuable guide for the analysis of human diseases.

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Peer Review #1 of "Identifying suitable reference genes for gene expression analysis in developing skeletal muscle in pigs (v0.1)"

10.7287/peerj.2428v0.1/reviews/1 ◽

2016 ◽

Keyword(s):

Gene Expression ◽

Skeletal Muscle ◽

Peer Review ◽

Expression Analysis ◽

Reference Genes ◽

Gene Expression Analysis ◽

Suitable Reference

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Peer Review #2 of "Identifying suitable reference genes for gene expression analysis in developing skeletal muscle in pigs (v0.1)"

10.7287/peerj.2428v0.1/reviews/2 ◽

2016 ◽

Keyword(s):

Gene Expression ◽

Skeletal Muscle ◽

Peer Review ◽

Expression Analysis ◽

Reference Genes ◽

Gene Expression Analysis ◽

Suitable Reference

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Selection of Suitable Reference Genes for RT-qPCR Gene Expression Analysis in Siberian Wild Rye (Elymus sibiricus) under Different Experimental Conditions

Genes ◽

10.3390/genes10060451 ◽

2019 ◽

Vol 10 (6) ◽

pp. 451 ◽

Cited By ~ 3

Author(s):

Junchao Zhang ◽

Wengang Xie ◽

Xinxuan Yu ◽

Zongyu Zhang ◽

Yongqiang Zhao ◽

...

Keyword(s):

Gene Expression ◽

Expression Analysis ◽

Reference Genes ◽

Gene Expression Analysis ◽

Expression Stability ◽

Forage Production ◽

Experimental Conditions ◽

Elymus Sibiricus ◽

Suitable Reference ◽

Selection Of

Elymus sibiricus, which is a perennial and self-pollinated grass, is the typical species of the genus Elymus, which plays an important role in forage production and ecological restoration. No reports have, so far, systematically described the selection of optimal reference genes for reverse transcriptase quantitative real-time polymerase chain reaction (RT-qPCR) analysis in E. sibiricus. The goals of this study were to evaluate the expression stability of 13 candidate reference genes in different experimental conditions, and to determine the appropriate reference genes for gene expression analysis in E. sibiricus. Five methods including Delta Ct (ΔCt), BestKeeper, NormFinder, geNorm, and RefFinder were used to assess the expression stability of 13 potential reference genes. The results of the RefFinder analysis showed that TBP2 and HIS3 were the most stable reference genes in different genotypes. TUA2 and PP2A had the most stable expression in different developmental stages. TBP2 and PP2A were suitable reference genes in different tissues. Under salt stress, ACT2 and TBP2 were identified as the most stable reference genes. ACT2 and TUA2 showed the most stability under heat stress. For cold stress, PP2A and ACT2 presented the highest degree of expression stability. DNAJ and U2AF were considered as the most stable reference genes under osmotic stress. The optimal reference genes were selected to investigate the expression pattern of target gene CSLE6 in different conditions. This study provides suitable reference genes for further gene expression analysis using RT-qPCR in E. sibiricus.

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Identification of Suitable Reference Genes for Quantitative Gene Expression Analysis in Innervated and Denervated Adipose Tissue from Cafeteria Diet‐Fed Rats

Lipids ◽

10.1002/lipd.12144 ◽

2019 ◽

Vol 54 (4) ◽

pp. 231-244 ◽

Cited By ~ 3

Author(s):

Gleuber Henrique Marques‐Oliveira ◽

Thaís Marques Silva ◽

Helder Magno Silva Valadares ◽

Helena Fonseca Raposo ◽

Ruither de Oliveira Gomes Carolino ◽

...

Keyword(s):

Gene Expression ◽

Adipose Tissue ◽

Expression Analysis ◽

Reference Genes ◽

Gene Expression Analysis ◽

Cafeteria Diet ◽

Quantitative Gene Expression ◽

Suitable Reference

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Identification of suitable reference genes for gene expression analysis of pork meat quality and analysis of candidate genes associated with the trait drip loss

Meat Science ◽

10.1016/j.meatsci.2010.05.030 ◽

2010 ◽

Vol 86 (2) ◽

pp. 436-439 ◽

Cited By ~ 17

Author(s):

Jean McBryan ◽

Ruth M. Hamill ◽

Grace Davey ◽

Peadar Lawlor ◽

Anne Maria Mullen

Keyword(s):

Gene Expression ◽

Candidate Genes ◽

Meat Quality ◽

Expression Analysis ◽

Reference Genes ◽

Gene Expression Analysis ◽

Drip Loss ◽

Pork Meat ◽

Suitable Reference

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Expression Stability of Internal Reference Gene in Response to Trichoderma polysporum Infection in Avena fatua L.

10.21203/rs.3.rs-507940/v1 ◽

2021 ◽

Author(s):

Haixia Zhu ◽

Yongqiang Ma ◽

Liang Cheng

Keyword(s):

Gene Expression ◽

Reference Gene ◽

Expression Analysis ◽

Reference Genes ◽

Gene Expression Analysis ◽

Avena Fatua ◽

Expression Stability ◽

Internal Reference ◽

Qpcr Analysis ◽

Suitable Combination

Abstract In order to construct a RT-qPCR system suitable for response of Avena fatua L. to Trichoderma polysporum , and screen stable internal reference genes, GeNorm, NormFinder, BestKeeper and RefFinde were used to perform SYBR Green-based RT-qPCR analysis on 8 candidate internal reference genes ( 18S , 28S , TUA , UBC , ACT , GAPDH , TBP and EF-1 ) in A. fatua samples after inoculation of T. polysporum Strain HZ-31. The results showed that TBP , 18S and UBC were the most stable internal reference genes, TBP and TUA , TBP and GAPDH , 18S and TBP , UBC and 18S were the most suitable combination of the two internal reference genes, which could be used as the internal reference genes for functional gene expression analysis during the interaction between T. polysporum and A. fatua .

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