Comprehensive selection of reference genes for quantitative RT-PCR analysis of murine extramedullary hematopoiesis during development

Medrano, Giuliana; Guan, Peihong; Barlow‐Anacker, Amanda J.; Gosain, Ankush

doi:10.1371/journal.pone.0181881

Cited by 22 publications

(12 citation statements)

References 36 publications

(35 reference statements)

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“…It was noteworthy though that Gapdh and B2m were identified as the most stable reference genes in the liver in the present study, as previous studies have also identified Gapdh [30] and B2m [31] as being the most stable reference genes in the liver in mouse models of obesity, with similar results also obtained by Matouskova et al [31]. Herein, Ywhaz and Gapdh were identified as the most stable reference genes in the spleen, which does not agree with results of previous studies in mouse models [32,33]. Ppia, Actb, Sdha, and B2m showed the greatest stability in the kidney tissue of hypercholesterolaemic rabbits, which supports a previous study of appropriate reference genes in kidney tissue that also found Actb to be one of the best reference genes [34].…”

Section: Discussioncontrasting

confidence: 88%

Evaluation of Reference Genes for Quantitative PCR in Four Tissues from Rabbits with Hypercholesterolaemia

Zhang

Wen

et al. 2019

Braz. arch. biol. technol.

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Rabbit with hypercholesterolaemia is an important model for studying cholesterol metabolism disease. This study aimed to evaluate the expression stability of nine reference genes for quantitative PCR (qPCR) analysis in adrenal gland, liver, spleen, and kidney tissue from rabbits with hypercholesterolaemia. In total, 30 male Harbin Large White (HLW) rabbits were fed a normal feed (n = 15) or a high cholesterol feed (n = 15) for 8 weeks to induce hypercholesterolaemia. Nine reference genes were verified by qPCR using cDNA extracted from rabbit tissue samples. For qPCR analysis, reference genes were evaluated using the RefFinder and GeNorm algorithms. Overall, seven rabbits with hypercholesterolaemia were identified based on body weight and total cholesterol measurements. Combining the results of the RefFinder and GeNorm algorithms, the most stable reference genes were hypoxanthine phosphoribosyltransferase 1 (Hprt1) and eukaryotic translation elongation factor 1 alpha 1 (Eef1a1) in the adrenal gland, β-2-microglobulin (B2m) and glyceraldehyde-3-phosphate dehydrogenase (Gapdh) in the liver, tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta (Ywhaz) and Gapdh in the spleen, and peptidylprolyl isomerase (Ppia), β-actin (Actb), succinate dehydrogenase complex subunit A flavoprotein (Sdha), and B2m in the kidney. Taken together, our results confirmed that Hprt1 and Eef1a1, B2m and Gapdh, Ywhaz and Gapdh, and Ppia, Actb, Sdha, and B2m were the best reference genes for qPCR analyses in adrenal gland, liver, spleen, and kidney tissue, respectively, of rabbits with hypercholesterolaemia.

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Section: Discussioncontrasting

confidence: 88%

Evaluation of Reference Genes for Quantitative PCR in Four Tissues from Rabbits with Hypercholesterolaemia

Zhang

Wen

et al. 2019

Braz. arch. biol. technol.

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“…BAG3 mRNA expression was analyzed using a real-time PCR instrument, TP800 (Takara, Japan). The relative BAG3 expression was normalized to 18S rRNA, and data analyses were performed using the comparative CT method [ 25 ].…”

Section: Methodsmentioning

confidence: 99%

Bcl-2-associated athanogene 3(BAG3) is associated with tumor cell proliferation, migration, invasion and chemoresistance in colorectal cancer

Chen

Cao

et al. 2018

BMC Cancer

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BackgroundCRC is one of the most common malignancies worldwide, and its molecular mechanisms remain unclear. Elevated levels of BAG3 have been reported in various tumors. The present study aimed to explore the expression and function of BAG3 in CRC.MethodsBAG3 protein expression was evaluated in 90 CRC specimens using immunohistochemistry in tissue microarrays, and the correlation between BAG3 expression and the clinicopathological features were assessed. In HCT116 cells BAG3 overexpression cell models were constructed, and CRISPR/Cas9 was used for BAG3 knockout. Western blotting and quantitative real-time PCR were used to determine BAG3 expression in HCT-116 Cells. Cell proliferation, migration and invasion were analyzed by cell counting, colony formation assay, EdU cell proliferation assay, RTCA growth curve assays, wound-healing migration assay and transwell invasion assay. The influence of BAG3 expression level on chemoresistance in HCT-116 cells was examined. Gene expression microarray and IPA analyses were employed to explore signaling pathways associated with the control of BAG3.ResultsUsing immunohistochemistry, this study found that BAG3 was markedly upregulated in colorectal cancer tissues and that BAG3 levels were significantly associated with tumor size and gender. BAG3 overexpression promoted HCT-116 cell growth, migration and invasion in vitro. In contrast, BAG3 knockout inhibited HCT-116 cell growth, migration and invasion. HCT-116 cells with high expression of BAG3 had higher cell viability and lower apoptosis rate than control cells after treatment with 5-FU, while the BAG3 knockout group demonstrated the opposite effects. So BAG3 expression level was associated with chemoresistance to 5-FU in HCT-116 cells. Gene expression microarrays and bioinformatics analyses of HCT-116 cells with BAG3 knockout demonstrated the involvement of BAG3 in signaling pathways associated with the control of cell proliferation, migration, invasion and chemoresistance in CRC.ConclusionsIn conclusion, this study provided evidence that BAG3 has a relevant role in CRC biology, and defined potential molecular pathways and networks. So BAG3 may be considered as a potential therapeutic target for anti-tumor therapy in colorectal cancer.

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“…Recently, Medrano and associates [13] demonstrated that, in the thymus, among six tested genes, Tbp (TATA box binding protein) gene was the most stable gene during the period, versus Hprt1 during the post-natal period (Day 21). Contrary to our current study, that investigation was performed on inbred mice (C57BL/6J strain) and only in the context of development, without addressing the infectious context.…”

Section: Discussionmentioning

confidence: 99%

Housekeeping Gene Expression in the Fetal and Neonatal Murine Thymus Following Coxsackievirus B4 Infection

Halouani

Jmii

Michaux

et al. 2020

Genes

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The thymus fulfills the role of T-cell production and differentiation. Studying transcription factors and genes involved in T-cell differentiation and maturation during the fetal and neonatal periods is very important. Nevertheless, no studies to date have been interested in evaluating the expressions of housekeeping genes as internal controls to assess the varying expressions of different genes inside this tissue during that period or in the context of viral infection. Thus, we evaluated by real-time quantitative polymerase chain reaction (qPCR) the expression of the most common internal control genes in the thymus of Swiss albino mice during the fetal and neonatal period, and following in utero infection with Coxsackievirus B4. The stability of expression of these reference genes in different samples was investigated using the geNorm application. Results demonstrated that the expression stability varied greatly between genes. Oaz1 was found to have the highest stability in different stages of development, as well as following Coxsackievirus B4 infection. The current study clearly demonstrated that Oaz1, with very stable expression levels that outperformed other tested housekeeping genes, could be used as a reference gene in the thymus and thymic epithelial cells during development and following Coxsackievirus B4 infection.

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Comprehensive selection of reference genes for quantitative RT-PCR analysis of murine extramedullary hematopoiesis during development

Cited by 22 publications

References 36 publications

Evaluation of Reference Genes for Quantitative PCR in Four Tissues from Rabbits with Hypercholesterolaemia

Evaluation of Reference Genes for Quantitative PCR in Four Tissues from Rabbits with Hypercholesterolaemia

Bcl-2-associated athanogene 3(BAG3) is associated with tumor cell proliferation, migration, invasion and chemoresistance in colorectal cancer

Housekeeping Gene Expression in the Fetal and Neonatal Murine Thymus Following Coxsackievirus B4 Infection

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