Expression Profiles of Glutathione S-Transferase Genes in Bombyx mori Exposed to NaF

2011 ◽  
Vol 175-176 ◽  
pp. 56-60
Author(s):  
Guo Dong Zhao ◽  
Yi Ling Zhang ◽  
Rui Xian Wang ◽  
Si Si Zhao ◽  
Ran Peng ◽  
...  
Keyword(s):  
Bombyx Mori ◽  
Real Time ◽  
Sodium Fluoride ◽  
Real Time Pcr ◽  
Reference Genes ◽  
Expression Profiles ◽  
Glutathione S Transferase ◽  
Internal Reference ◽  
Pcr Method ◽  
Different Tissues

In order to explore the roles of Bombyx mori glutathione S-transferase gene (BmGST) in detoxification and resistance to insecticides, we used real-time PCR method to detect the transcription levels of five BmGST genes in different tissues of the 5th instar larvae feeding on sodium fluoride treated mulberry leaves. The detection results were normalized by using 3 internal reference genes. The results indicated that the transcription levels of BmGSTs were different in various tissues. Transcription of BmGST genes could be induced by NaF, The normalized data with the above 3 internal reference genes indicated that, it is very important to choose adequate internal reference genes so as to ensure the reliability of the detection results.

scholarly journals Cloning and evaluation of reference genes for quantitative real-time PCR analysis inAmorphophallus

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3260 ◽  
Author(s):  
Kai Wang ◽  
Yi Niu ◽  
Qijun Wang ◽  
Haili Liu ◽  
Yi Jin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heat Stress ◽  
Real Time ◽  
Reference Genes ◽  
Dynamic Range ◽  
Expression Profiles ◽  
Mrna Levels ◽  
Expression Stability ◽  
Degenerate Primers ◽  
Different Tissues

Quantitative real-time reverse transcription PCR (RT-qPCR) has been widely used in the detection and quantification of gene expression levels because of its high accuracy, sensitivity, and reproducibility as well as its large dynamic range. However, the reliability and accuracy of RT-qPCR depends on accurate transcript normalization using stably expressed reference genes.Amorphophallusis a perennial plant with a high content of konjac glucomannan (KGM) in its corm. This crop has been used as a food source and as a traditional medicine for thousands of years. Without adequate knowledge of gene expression profiles, there has been no report of validated reference genes inAmorphophallus. In this study, nine genes that are usually used as reference genes in other crops were selected as candidate reference genes. These putative sequences of these genesAmorphophalluswere cloned by the use of degenerate primers. The expression stability of each gene was assessed in different tissues and under two abiotic stresses (heat and waterlogging) inA. albusandA. konjac. Three distinct algorithms were used to evaluate the expression stability of the candidate reference genes. The results demonstrated thatEF1-a,EIF4A,H3andUBQwere the best reference genes under heat stress inAmorphophallus. Furthermore,EF1-a,EIF4A,TUB, andRPwere the best reference genes in waterlogged conditions. By comparing different tissues from all samples, we determined thatEF1-α,EIF4A,andCYPwere stable in these sets. In addition, the suitability of these reference genes was confirmed by validating the expression of a gene encoding the small heat shock proteinSHSP, which is related to heat stress inAmorphophallus. In sum,EF1-αandEIF4Awere the two best reference genes for normalizing mRNA levels in different tissues and under various stress treatments, and we suggest using one of these genes in combination with 1 or 2 reference genes associated with different biological processes to normalize gene expression. Our results will provide researchers with appropriate reference genes for further gene expression quantification using RT-qPCR inAmorphophallus.

scholarly journals Evaluation of Internal Reference Genes for Quantitative Expression Analysis by Real-Time PCR in Ovine Whole Blood

2011 ◽  
Vol 12 (11) ◽  
pp. 7732-7747 ◽  
Author(s):  
Simone Peletto ◽  
Simone Bertuzzi ◽  
Chiara Campanella ◽  
Paola Modesto ◽  
Maria Grazia Maniaci ◽  
...  
Keyword(s):  
Real Time ◽  
Expression Analysis ◽  
Real Time Pcr ◽  
Whole Blood ◽  
Reference Genes ◽  
Internal Reference ◽  
Quantitative Expression

scholarly journals Selection and Validation of Reference Genes for Quantitative Real-Time PCR Expression Analysis of Candidate Genes in Carposina sasakii (Lepidoptera: Carposinidae)

2020 ◽  
pp. 75-85
Author(s):  
Zuobing Yan ◽  
Yongli Li ◽  
Zhou Zhou ◽  
Yongan Zhang ◽  
Liangjian Qu
Keyword(s):  
Gene Expression ◽  
Real Time ◽  
Expression Analysis ◽  
Real Time Pcr ◽  
Reference Genes ◽  
Developmental Stages ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Quantitative Real Time Pcr ◽  
Carposina Sasakii

Carposina sasakii is one of the most important pests on the quality of stone and pome fruits. Investigation of a gene expression level in the species is hampered because of the gap of validated reference genes. The expression variation in the transcription levels of eight candidate reference genes, Actin (ACT), Tubulinbeta-1 (TUB), Ribosomal protein 49 (RP49), Elongation factor1-alpha (EF-1a), Elongation factor1-b (EF-1b), Elongation factor1-d (EF-1d), Ribosomal proteinL13 (RPL13) and Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), were analyzed by quantitative real-time PCR (qPCR). The stability and ranking of these gene expression profiles in three organ types (head, thorax and abdomen), three developmental stages (larva, pupa and moth), and five diapause states (non-diapause, pre-diapause, diapause 0 d, diapause 20 d and diapause 60 d) were assessed using two algorithm-based methods, geNorm and NormFinder. EF-1a, ACT and GAPDH were evaluated to be the three stable reference genes based on the important observations and comprehensive analysis, whereas TUB and EF-1b showed low expression stability. Best gene combinations for different qPCR analysis in C. sasakii could be chosen from the three stable reference genes, the using of two reference genes is sufficient to effectively normalize qPCR data in C. sasakii. The study laid the foundation for gene expression analysis in C. sasakii and provided new information for the selection of reference genes.

Selection and validation of reference genes for reverse transcription quantitative real-time PCR (RT-qPCR) in silkworm infected with Bombyx mori bidensovirus

Biologia ◽  
2018 ◽  
Vol 73 (9) ◽  
pp. 897-906 ◽  
Author(s):  
Zhaoyang Hu ◽  
Yanchun Deng ◽  
Xiaolong Zhang ◽  
Peipei Tang ◽  
Weijuan Sun ◽  
...  
Keyword(s):  
Bombyx Mori ◽  
Real Time ◽  
Reverse Transcription ◽  
Real Time Pcr ◽  
Reference Genes ◽  
Quantitative Real Time Pcr

scholarly journals Tissue-specific selection of stable reference genes for real-time PCR normalization in an obese rat model

2012 ◽  
Vol 48 (3) ◽  
pp. 251-260 ◽  
Author(s):  
Manuela Cabiati ◽  
Serena Raucci ◽  
Chiara Caselli ◽  
Maria Angela Guzzardi ◽  
Andrea D'Amico ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Real Time ◽  
Rat Model ◽  
Real Time Pcr ◽  
Reference Genes ◽  
Genetic Model ◽  
Expression Profiles ◽  
Zucker Rats ◽  
Specific Reference ◽  
Zucker Rat

Obesity is a complex pathology with interacting and confounding causes due to the environment, hormonal signaling patterns, and genetic predisposition. At present, the Zucker rat is an eligible genetic model for research on obesity and metabolic syndrome, allowing scrutiny of gene expression profiles. Real-time PCR is the benchmark method for measuring mRNA expressions, but the accuracy and reproducibility of its data greatly depend on appropriate normalization strategies. In the Zucker rat model, no specific reference genes have been identified in myocardium, kidney, and lung, the main organs involved in this syndrome. The aim of this study was to select among ten candidates (Actb, Gapdh, Polr2a, Ywhag, Rpl13a, Sdha, Ppia, Tbp, Hprt1andTfrc) a set of reference genes that can be used for the normalization of mRNA expression data obtained by real-time PCR in obese and lean Zucker rats both at fasting and during acute hyperglycemia. The most stable genes in the heart wereSdha, Tbp, andHprt1; in kidney,Tbp,Actb, andGapdhwere chosen, whileActb,Ywhag, andSdhawere selected as the most stably expressed set for pulmonary tissue. The normalization strategy was used to analyze mRNA expression of tumor necrosis factor α, the main inflammatory mediator in obesity, whose variations were more significant when normalized with the appropriately selected reference genes. The findings obtained in this study underline the importance of having three stably expressed reference gene sets for use in the cardiac, renal, and pulmonary tissues of an experimental model of obese and hyperglycemic Zucker rats.

scholarly journals Identification and evaluation of reference genes for quantitative real-time PCR analysis in Polygonum cuspidatum based on transcriptome data

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Xiaowei Wang ◽  
Zhijun Wu ◽  
Wenqi Bao ◽  
Hongyan Hu ◽  
Mo Chen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Real Time ◽  
Stress Responses ◽  
Real Time Pcr ◽  
Abiotic Stresses ◽  
Reference Genes ◽  
Expression Patterns ◽  
Polygonum Cuspidatum ◽  
Quantitative Real Time Pcr ◽  
Different Tissues

Abstract Background Polygonum cuspidatum of the Polygonaceae family is a traditional medicinal plant with many bioactive compounds that play important roles in human health and stress responses. Research has attempted to identify biosynthesis genes and metabolic pathways in this species, and quantitative real-time PCR (RT-qPCR) has commonly been used to detect gene expression because of its speed, sensitivity, and specificity. However, no P. cuspidatum reference genes have been identified, which hinders gene expression studies. Here, we aimed to identify suitable reference genes for accurate and reliable normalization of P. cuspidatum RT-qPCR data. Results Twelve candidate reference genes, including nine common (ACT, TUA, TUB, GAPDH, EF-1γ, UBQ, UBC, 60SrRNA, and eIF6A) and three novel (SKD1, YLS8, and NDUFA13), were analyzed in different tissues (root, stem, and leaf) without treatment and in leaves under abiotic stresses (salt, ultraviolet [UV], cold, heat, and drought) and hormone stimuli (abscisic acid [ABA], ethylene [ETH], gibberellin [GA3], methyl jasmonate [MeJA], and salicylic acid [SA]). Expression stability in 65 samples was calculated using the △CT method, geNorm, NormFinder, BestKeeper, and RefFinder. Two reference genes (NDUFA13 and EF-1γ) were sufficient to normalize gene expression across all sample sets. They were also the two most stable genes for abiotic stresses and different tissues, whereas NDUFA13 and SKD1 were the top two choices for hormone stimuli. Considering individual experimental sets, GAPDH was the top-ranked gene under ABA, ETH, and GA3 treatments, while 60SrRNA showed good stability under MeJA and cold treatments. ACT, UBC, and TUB were suitable genes for drought, UV, and ABA treatments, respectively. TUA was not suitable because of its considerable variation in expression under different conditions. The expression patterns of PcPAL, PcSTS, and PcMYB4 under UV and SA treatments and in different tissues normalized by stable and unstable reference genes demonstrated the suitability of the optimal reference genes. Conclusions We propose NDUFA13 and EF-1γ as reference genes to normalize P. cuspidatum expression data. To our knowledge, this is the first systematic study of reference genes in P. cuspidatum which could help advance molecular biology research in P. cuspidatum and allied species.

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