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

Genes ◽  
2019 ◽  
Vol 10 (6) ◽  
pp. 451 ◽  
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.

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

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2428 ◽  
Author(s):  
Guanglin Niu ◽  
Yalan Yang ◽  
YuanYuan Zhang ◽  
Chaoju Hua ◽  
Zishuai Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Expression Analysis ◽  
Reference Genes ◽  
Gene Expression Analysis ◽  
Muscle Development ◽  
Expression Stability ◽  
Skeletal Muscle Development ◽  
Expression Levels ◽  
Suitable Reference

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.

scholarly journals Validation of Internal Control Genes for Quantitative Real-Time PCR Gene Expression Analysis in Morchella

Molecules ◽  
2018 ◽  
Vol 23 (9) ◽  
pp. 2331 ◽  
Author(s):  
Qianqian Zhang ◽  
Wei Liu ◽  
Yingli Cai ◽  
A-Feng Lan ◽  
Yinbing Bian
Keyword(s):  
Gene Expression ◽  
Candidate Genes ◽  
Expression Analysis ◽  
Internal Control ◽  
Reference Genes ◽  
Gene Expression Analysis ◽  
Stable Gene ◽  
Experimental Conditions ◽  
The Stability ◽  
Vacuolar Protein

The reliability of qRT-PCR results depend on the stability of reference genes used for normalization, suggesting the necessity of identification of reference genes before gene expression analysis. Morels are edible mushrooms well-known across the world and highly prized by many culinary kitchens. Here, several candidate genes were selected and designed according to the Morchella importuna transcriptome data. The stability of the candidate genes was evaluated with geNorm and NormFinder under three different experimental conditions, and several genes with excellent stability were selected. The extensive adaptability of the selected genes was tested in ten Morchella species. Results from the three experimental conditions revealed that ACT1 and INTF7 were the most prominent genes in Morchella, CYC3 was the most stable gene in different development stages, INTF4/AEF3 were the top-ranked genes across carbon sources, while INTF3/CYC3 pair showed the robust stability for temperature stress treatment. We suggest using ACT1, AEF3, CYC3, INTF3, INTF4 and INTF7 as reference genes for gene expression analysis studies for any of the 10 Morchella strains tested in this study. The stability and practicality of the gene, vacuolar protein sorting (INTF3), vacuolar ATP synthase (INTF4) and14-3-3 protein (INTF7) involving the basic biological processes were validated for the first time as the candidate reference genes for quantitative PCR. Furthermore, the stability of the reference genes was found to vary under the three different experimental conditions, indicating the importance of identifying specific reference genes for particular conditions.

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

Lipids ◽  
2019 ◽  
Vol 54 (4) ◽  
pp. 231-244 ◽  
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

scholarly journals Genome Based Search for Reference Genes for Gene Expression Analysis in Oral Cancer: a Data Science Driven Approach

2019 ◽  
Author(s):  
Nehanjali Dwivedi ◽  
Sujan K Dhar ◽  
G Charitha ◽  
Moni Abraham Kuriakose ◽  
Amritha Suresh ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Expression Analysis ◽  
Reference Genes ◽  
Data Science ◽  
Gene Expression Analysis ◽  
The Cancer Genome Atlas ◽  
Experimental Conditions ◽  
Statistical Measures ◽  
Different Types

Abstract Background Quantitative real time PCR (qPCR) remains by far the most cost-effective, fast yet sensitive technique to check the gene expression levels in various systems. The traditionally used reference genes over the years were found to be regulated heavily based on sample sources and/or experimental conditions. This paper therefore presents a data science driven -omic approach for selection of reference genes from ~60,000 candidates from The Cancer Genome Atlas (TCGA) and Broad Institute Cancer Cell Line Encyclopaedia (CCLE) for gene expression studies in head and neck squamous cell carcinoma (HNSCC). mRNA-sequencing data of 500 patient samples and 33 cell lines from publicly available databases were analysed to assess stability of genes in terms of multiple statistical measures. A final set of 12 candidate genes were studied in the Indian set of data in Gene Expression Omnibus (GEO) and validated experimentally using qPCR in 35 different types of samples from platforms like drug sensitive and resistant cell lines, normal and tumor samples, fibroblast and epithelial primary culture derived from HNSCC patients from India. Result The study lead to the choice of five most stable reference genes –TYW5, RIC8B, PLEKHA3, CEP57L1 and GPR89B across three experimental platforms. Conclusion In addition to providing a set of five most stable reference genes for future gene expression analysis experiments across different types of samples in HNSCC, the study also presents an objective framework for assessing reference genes for other disease areas as well.

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

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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