Housekeeping Genes in Cancer: Normalization of Array Data

Khimani, Anis H.; Mhashilkar, Abner M.; Mikulskis, Alvydas; O’Malley, Michael S.; Liao, Jennifer; Golenko, Eva; Mayer, Pat; Chada, Sunil; Killian, Jeffrey; Lott, Steven T.

doi:10.2144/05385st04

Cited by 59 publications

(44 citation statements)

References 32 publications

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“…Overall, the majority of these mRNAs are effective normalization tools, although several groups report that variability in their expression levels can exist in different sample and cell types and needs to be considered when designing a study. [1][2][3][4][5][6][7][8] For example, Vandesompele et al 9 reported that universally stable control genes do not exist, thus necessitating the development of an accurate normalization strategy for each experimental system. Aerts et al, 10 in a study of 26 tumor cell lines including prostate cancer DU-145 and PC-3, showed GAPDH has the highest variability in gene expression of commonly used controls, whereas 18s rRNA, GUS, and ACTB had the best expression stability.…”

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confidence: 99%

Assessment of normalization strategies for quantitative RT-PCR using microdissected tissue samples

Erickson

Albert

Gillespie

et al. 2007

Laboratory Investigation

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Gene expression measurement techniques such as quantitative reverse transcriptase (qRT)-PCR require a normalization strategy to allow meaningful comparisons across biological samples. Typically, this is accomplished through the use of an endogenous housekeeping gene that is presumed to show stable expression levels in the samples under study. There is concern regarding how precisely specific genes can be measured in limited amounts of mRNA such as those from microdissected (MD) tissues. To address this issue, we evaluated three different approaches for qRT-PCR normalization of dissected samples; cell count during microdissection, total RNA measurement, and endogenous control genes. The data indicate that both cell count and total RNA are useful in calibrating input amounts at the outset of a study, but do not provide enough precision to serve as normalization standards. However, endogenous control genes can accurately determine the relative abundance of a target gene relative to the entire cellular transcriptome. Taken together, these results suggest that precise gene expression measurements can be made from MD samples if the appropriate normalization strategy is employed.

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mentioning

confidence: 99%

Assessment of normalization strategies for quantitative RT-PCR using microdissected tissue samples

Erickson

Albert

Gillespie

et al. 2007

Laboratory Investigation

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“…This protein has been considered a housekeeping enzyme. However, it has been reported dysregulated in cancer cells [51], liver metastases [52], and in aged rats [53]. In COMT deleted mice, the changes in actin 1 might also be related to the response to stress in liver.…”

Section: Proteome Of Female Comt Knockout Livermentioning

confidence: 99%

Sex-dependent compensated oxidative stress in the mouse liver upon deletion of catechol O-methyltransferase

Tenorio-Laranga¹,

Männistö²,

Karayiorgou³

et al. 2009

Biochemical Pharmacology

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. Sex-dependent compensated oxidative stress in the mouse liver upon deletion of catechol O-methyltransferase. Biochemical Pharmacology, Elsevier, 2009, 77 (9) This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.Page 1 of 50 A c c e p t e d M a n u s c r i p

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“…An extensive literature has discussed problems related to normalization in such studies. One such problem is a change in expression of the gene/protein used for normalization-a particular concern in studies of development, differentiation, or disease (Khimani et al, 2005;Brattelid et al, 2007). Two issues in the use of arrays to assess GPCR expression are how to define the limit of detection of a receptor in terms of Δcycle-threshold [ΔC(t)] relative to the reference used for normalization, and the most appropriate statistical tests for data analysis (Khimani et al, 2005;Rubie et al, 2005).…”

Section: What Are Some Of the Problems And Limitations Of Efforts To mentioning

confidence: 99%

G Protein–Coupled Receptor (GPCR) Expression in Native Cells: “Novel” endoGPCRs as Physiologic Regulators and Therapeutic Targets

et al. 2015

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G protein-coupled receptors (GPCRs), the largest family of signaling receptors in the human genome, are also the largest class of targets of approved drugs. Are the optimal GPCRs (in terms of efficacy and safety) currently targeted therapeutically? Especially given the large number (∼120) of orphan GPCRs (which lack known physiologic agonists), it is likely that previously unrecognized GPCRs, especially orphan receptors, regulate cell function and can be therapeutic targets. Knowledge is limited regarding the diversity and identity of GPCRs that are activated by endogenous ligands and that native cells express. Here, we review approaches to define GPCR expression in tissues and cells and results from studies using these approaches. We identify problems with the available data and suggest future ways to identify and validate the physiologic and therapeutic roles of previously unrecognized GPCRs. We propose that a particularly useful approach to identify functionally important GPCRs with therapeutic potential will be to focus on receptors that show selective increases in expression in diseased cells from patients and experimental animals.

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Housekeeping Genes in Cancer: Normalization of Array Data

Cited by 59 publications

References 32 publications

Assessment of normalization strategies for quantitative RT-PCR using microdissected tissue samples

Assessment of normalization strategies for quantitative RT-PCR using microdissected tissue samples

Sex-dependent compensated oxidative stress in the mouse liver upon deletion of catechol O-methyltransferase

G Protein–Coupled Receptor (GPCR) Expression in Native Cells: “Novel” endoGPCRs as Physiologic Regulators and Therapeutic Targets

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