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Normfinder injury

Tianyi Li, Hongying Diao, Lei Zhao, Yue Xing, Jichang Zhang, Ning Liu, Youyou Yan, Xin Tian, Wei Sun, Bin Liu
BACKGROUND: Oxidative stress can induce cell injury in vascular endothelial cells, which is the initial event in the development of atherosclerosis. Although quantitative real-time polymerase chain reaction (qRT-PCR) has been widely used in gene expression studies in oxidative stress injuries, using carefully validated reference genes has not received sufficient attention in related studies. The objective of this study, therefore, was to select a set of stably expressed reference genes for use in qRT-PCR normalization in oxidative stress injuries in human umbilical vein endothelial cells (HUVECs) induced by hydrogen peroxide (H2O2)...
April 5, 2017: BMC Molecular Biology
Prue M Pereira-Fantini, Anushi E Rajapaksa, Regina Oakley, David G Tingay
Preterm newborns often require invasive support, however even brief periods of supported ventilation applied inappropriately to the lung can cause injury. Real-time quantitative reverse transcriptase-PCR (qPCR) has been extensively employed in studies of ventilation-induced lung injury with the reference gene 18S ribosomal RNA (18S RNA) most commonly employed as the internal control reference gene. Whilst the results of these studies depend on the stability of the reference gene employed, the use of 18S RNA has not been validated...
2016: Scientific Reports
Mariana Ferreira Leal, Gustavo Gonçalves Arliani, Diego Costa Astur, Carlos Eduardo Franciozi, Pedro Debieux, Carlos Vicente Andreoli, Marília Cardoso Smith, Alberto de Castro Pochini, Benno Ejnisman, Moises Cohen
The meniscus plays critical roles in the knee function. Meniscal tears can lead to knee osteoarthritis. Gene expression analysis may be a useful tool for understanding meniscus tears, and reverse-transcription quantitative polymerase chain reaction (RT-qPCR) has become an effective method for such studies. However, this technique requires the use of suitable reference genes for data normalization. We evaluated the suitability of six reference genes (18S, ACTB, B2M, GAPDH, HPRT1 and TBP) using meniscus samples of (1) 19 patients with isolated meniscal tears, (2) 20 patients with meniscal tears and combined anterior cruciate ligament injury (ACL), and (3) 11 controls without meniscal tears...
June 10, 2016: Gene
Yuting Yang, Xu Zhang, Yun Chen, Jinlong Guo, Hui Ling, Shiwu Gao, Yachun Su, Youxiong Que, Liping Xu
Sugarcane, accounting for 80% of world's sugar, originates in the tropics but is cultivated mainly in the subtropics. Therefore, chilling injury frequently occurs and results in serious losses. Recent studies in various plant species have established microRNAs as key elements in the post-transcriptional regulation of response to biotic and abiotic stresses including cold stress. Though, its accuracy is largely influenced by the use of reference gene for normalization, quantitative PCR is undoubtedly a popular method used for identification of microRNAs...
2016: Frontiers in Plant Science
Mariana Ferreira Leal, Diego Costa Astur, Pedro Debieux, Gustavo Gonçalves Arliani, Carlos Eduardo Silveira Franciozi, Leonor Casilla Loyola, Carlos Vicente Andreoli, Marília Cardoso Smith, Alberto de Castro Pochini, Benno Ejnisman, Moises Cohen
The anterior cruciate ligament (ACL) is one of the most frequently injured structures during high-impact sporting activities. Gene expression analysis may be a useful tool for understanding ACL tears and healing failure. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) has emerged as an effective method for such studies. However, this technique requires the use of suitable reference genes for data normalization. Here, we evaluated the suitability of six reference genes (18S, ACTB, B2M, GAPDH, HPRT1, and TBP) by using ACL samples of 39 individuals with ACL tears (20 with isolated ACL tears and 19 with ACL tear and combined meniscal injury) and of 13 controls...
2015: PloS One
Yu-Xi He, Yan Zhang, Qiwei Yang, Chenguang Wang, Guanfang Su
Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is a technique widely used for the quantification of mRNA transcription, It is well recognized that the reference genes used in RT-qPCR require appropriate validation to ensure that gene expression is unaffected by experimental conditions. The differentiation of bone mesenchymal stem cells (BMSCs) into neurons is important in the treatment of nerve injury. In gene expression analysis of the differentiation of BMSCs into neuronal cells by, the commonly used reference genes for RNA analysis are often selected without any preliminary evaluation of their suitability...
August 2015: Molecular Medicine Reports
Mariana Ferreira Leal, Paulo Santoro Belangero, Eduardo Antônio Figueiredo, Carina Cohen, Leonor Casilla Loyola, Carlos Vicente Andreoli, Marília Cardoso Smith, Alberto de Castro Pochini, Benno Ejnisman, Moises Cohen
Rotator cuff tear is one of the most common causes of shoulder dysfunction. Gene expression analysis may be a useful tool for understanding tendon tears and the failure of cuff healing, and reverse-transcription quantitative polymerase chain reaction (RT-qPCR) has become an effective method for such studies. However, this technique requires the use of suitable reference genes for data normalization. Here, we evaluate the suitability of six reference genes (18S, ACTB, B2M, GAPDH, HPRT1 and TBP) using samples from the rotator cuff tendons of 28 individuals with tendon tears (3 tendons regions) and 8 controls (2 tendon regions); for the tear patients, we evaluated ruptured and non-ruptured tendon samples...
2015: PloS One
Roxanne Y Walder, Anne-Sophie Wattiez, Stephanie R White, Blanca Marquez de Prado, Marta V Hamity, Donna L Hammond
BACKGROUND: Real-time quantitative PCR (qPCR) is a technique frequently used to measure changes in mRNA expression. To ensure validity of experimental findings, it is important to normalize the qPCR data to reference genes that are stable and unaffected by the experimental treatment to correct for variability among samples. Unlike in some models of neuropathic pain, reference genes for models of inflammatory injury have not been validated. This study examined four candidate reference genes in an effort to identify and validate optimal genes for normalization of transcriptional changes occurring in the dorsal horn of the spinal cord and the rostral ventromedial medulla (RVM) following intraplantar injection of complete Freund's adjuvant (CFA)...
2014: Molecular Pain
Giovanna Gambarotta, Giulia Ronchi, Olivier Friard, Pantaleo Galletta, Isabelle Perroteau, Stefano Geuna
Injury to the peripheral nerve induces dramatic changes in terms of cellular composition that are reflected on RNA quality and quantity, making messenger RNA expression analysis very complex. Several commonly used housekeeping genes are regulated following peripheral nerve injury and are thus not suitable for quantitative real-time PCR normalization; moreover, the presence of pseudogenes in some of them impairs their use. To deal with this problem, we have developed a new method to identify new stable housekeeping genes based on publicly available microarray data on normal and injured nerves...
2014: PloS One
Mariana Ferreira Leal, Paulo Santoro Belangero, Carina Cohen, Eduardo Antônio Figueiredo, Leonor Casilla Loyola, Alberto Castro Pochini, Marília Cardoso Smith, Carlos Vicente Andreoli, Sintia Iole Belangero, Benno Ejnisman, Moises Cohen
Shoulder instability is a common shoulder injury, and patients present with plastic deformation of the glenohumeral capsule. Gene expression analysis may be a useful tool for increasing the general understanding of capsule deformation, and reverse-transcription quantitative polymerase chain reaction (RT-qPCR) has become an effective method for such studies. Although RT-qPCR is highly sensitive and specific, it requires the use of suitable reference genes for data normalization to guarantee meaningful and reproducible results...
2014: PloS One
Wei Fu, Wen Xie, Zhuo Zhang, Shaoli Wang, Qingjun Wu, Yong Liu, Xiaomao Zhou, Xuguo Zhou, Youjun Zhang
Quantitative real-time PCR (qRT-PCR), a primary tool in gene expression analysis, requires an appropriate normalization strategy to control for variation among samples. The best option is to compare the mRNA level of a target gene with that of reference gene(s) whose expression level is stable across various experimental conditions. In this study, expression profiles of eight candidate reference genes from the diamondback moth, Plutella xylostella, were evaluated under diverse experimental conditions. RefFinder, a web-based analysis tool, integrates four major computational programs including geNorm, Normfinder, BestKeeper, and the comparative ΔCt method to comprehensively rank the tested candidate genes...
2013: International Journal of Biological Sciences
Yan Wang, Naping Tang, Taotao Hui, Shuyan Wang, Xiancheng Zeng, Hua Li, Jing Ma
Circulating microRNA (miRNA) expression profiles have been reported to be promising biomarkers for drug-induced liver injury in preclinical and clinical practice. Proper normalization is critical for accurate miRNAs expression analysis. Herein, using SYBR green quantitative real-time PCR (RT-qPCR), we evaluated the expression stability of six candidate reference genes including two commonly used small RNAs (U6, 5S) and four miRNAs (let-7a, miR-92a, miR-103 and miR-16) in plasma of rats with acetaminophen-induced hepatotoxicity...
November 2013: Journal of Applied Toxicology: JAT
Xiao-Xiang Peng, Rong-Lan Zhao, Wei Song, Hai-Rong Chu, Meng Li, Shu-Ya Song, Guang-Zhou Li, Dong-Chun Liang
When studying the altered expression of genes associated with cartilage regeneration by quantitative real-time RT-PCR (RT-qPCR), reference genes with highly stable expression during different stages of chondrocyte developmental are necessary to normalize gene expression accurately. Until now, no reports evaluating expression changes of commonly used reference genes in rabbit articular cartilage have been published. In this study, defects were made in rabbit articular cartilage, with or without insulin-like growth factor 1 (IGF-1) treatment, to create different chondrocyte living environments...
2012: International Journal of Molecular Sciences
Ryan S McCulloch, Melissa S Ashwell, Audrey T O'Nan, Peter L Mente
UNLABELLED: HASH(0x49912f0) BACKGROUND: Expression levels for genes of interest must be normalized with an appropriate reference, or housekeeping gene, to make accurate comparisons of quantitative real-time PCR results. The purpose of this study was to identify the most stable housekeeping genes in porcine articular cartilage subjected to a mechanical injury from a panel of 10 candidate genes. RESULTS: Ten candidate housekeeping genes were evaluated in three different treatment groups of mechanically impacted porcine articular cartilage...
2012: Journal of Animal Science and Biotechnology
Qi Wang, Takaki Ishikawa, Tomomi Michiue, Bao-Li Zhu, Da-Wei Guan, Hitoshi Maeda
In forensic molecular pathology, quantitative real-time polymerase chain reaction (RT-qPCR) provides a rapid and sensitive method to investigate functional changes in the death process. Accurate and reliable relative RT-qPCR requires ideal amplification efficiencies of target and reference genes. However, the amplification efficiency, changing during PCR, may be overestimated by the traditional standard curve method. No single gene meets the criteria of an ideal endogenous reference. Therefore, it is necessary to select suitable reference genes for specific requirements...
November 2012: International Journal of Legal Medicine
María N Lardizábal, Ana L Nocito, Stella M Daniele, Leonardo A Ornella, Javier F Palatnik, Luis M Veggi
Hepatotoxicity is associated with major changes in liver gene expression induced by xenobiotic exposure. Understanding the underlying mechanisms is critical for its clinical diagnosis and treatment. MicroRNAs are key regulators of gene expression that control mRNA stability and translation, during normal development and pathology. The canonical technique to measure gene transcript levels is Real-Time qPCR, which has been successfully modified to determine the levels of microRNAs as well. However, in order to obtain accurate data in a multi-step method like RT-qPCR, the normalization with endogenous, stably expressed reference genes is mandatory...
2012: PloS One
Madhavi Latha Yadav Bangaru, Frank Park, Andy Hudmon, J Bruce McCallum, Quinn H Hogan
Stably expressed housekeeping genes (HKGs) are necessary for standardization of transcript measurement by quantitative real-time polymerase chain reaction (qRT-PCR). Peripheral nerve injury disrupts expression of numerous genes in sensory neurons, but the stability of conventional HKGs has not been tested in this context. We examined the stability of candidate HKGs during nerve injury, including the commonly used 18S ribosomal RNA, β-tubulin I and β-tubulin III, actin, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and hypoxanthine phosphoribosyltransferase 1 (HPRT1), and mitogen-activated protein kinase 6 (MAPK6)...
March 2012: Journal of Molecular Neuroscience: MN
Shengwei Ren, Feng Zhang, Changyou Li, Changkai Jia, Siyuan Li, Haijie Xi, Hongbo Zhang, Lingling Yang, Yiqiang Wang
PURPOSE: To evaluate the suitability of common housekeeping genes (HKGs) for use in quantitative reverse transcription PCR (qRT-PCR) assays of the cornea in various murine disease models. METHODS: CORNEAL DISEASE MODELS STUDIED WERE: 1) corneal neovascularization (CorNV) induced by suture or chemical burn, 2) corneal infection with Candida albicans or Aspergillus fumigatus by intrastromal injection of live spores, and 3) perforating corneal injury (PCI) in Balb/c mice or C57BL/6 mice...
2010: Molecular Vision
Guojun Shi, Zhijian Zhang, Dechun Feng, Yan Xu, Yan Lu, Jiqiu Wang, Jingjing Jiang, Zhiguo Zhang, Xiaoying Li, Guang Ning
Quantitative real-time reverse transcription-polymerase chain reaction (Q-PCR) has become an indispensable technique for accurate determination of gene expression in various samples. In mice, intravenous injection of concanavalin A (ConA) leads to acute hepatitis and liver injury. Functional studies based on this model have provided insights for understanding the mechanisms of liver injury. However, no data have been reported to validate reference genes during the progression of ConA-induced hepatitis (CIH)...
June 1, 2010: Analytical Biochemistry
Hervé Rhinn, Catherine Marchand-Leroux, Nicole Croci, Michel Plotkine, Daniel Scherman, Virginie Escriou
BACKGROUND: Traumatic brain injury models are widely studied, especially through gene expression, either to further understand implied biological mechanisms or to assess the efficiency of potential therapies. A large number of biological pathways are affected in brain trauma models, whose elucidation might greatly benefit from transcriptomic studies. However the suitability of reference genes needed for quantitative RT-PCR experiments is missing for these models. RESULTS: We have compared five potential reference genes as well as total cDNA level monitored using Oligreen reagent in order to determine the best normalizing factors for quantitative RT-PCR expression studies in the early phase (0-48 h post-trauma (PT)) of a murine model of diffuse brain injury...
2008: BMC Molecular Biology
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