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https://www.readbyqxmd.com/read/28815002/a-comprehensive-method-for-identification-of-suitable-reference-genes-in-extracellular-vesicles
#1
Kenneth Gouin, Kiel Peck, Travis Antes, Jennifer Leigh Johnson, Chang Li, Sharon Denise Vaturi, Ryan Middleton, Geoff de Couto, Ann-Sophie Walravens, Luis Rodriguez-Borlado, Rachel Ruckdeschel Smith, Linda Marbán, Eduardo Marbán, Ahmed Gamal-Eldin Ibrahim
Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is one of the most sensitive, economical and widely used methods for evaluating gene expression. However, the utility of this method continues to be undermined by a number of challenges including normalization using appropriate reference genes. The need to develop tailored and effective strategies is further underscored by the burgeoning field of extracellular vesicle (EV) biology. EVs contain unique signatures of small RNAs including microRNAs (miRs)...
2017: Journal of Extracellular Vesicles
https://www.readbyqxmd.com/read/28810828/suitable-reference-gene-for-quantitative-real-time-pcr-analysis-of-gene-expression-in-gonadal-tissues-of-minnow-puntius-sophore-under-high-temperature-stress
#2
Arabinda Mahanty, Gopal Krishna Purohit, Sasmita Mohanty, Nihar Ranjan Nayak, Bimal Prasanna Mohanty
BACKGROUND: High ambient temperature is known to affect fish gonadal development and physiology in a variety of ways depending on the severity and duration of exposure; however, the underlying molecular mechanisms are poorly understood. Gonadal gene expression influence the gonadal development, physiology and the quality of egg/sperm produced in teleosts and the mechanistic understanding of spatio-temporal changes in the gonadal gene expression could be instrumental in controlling the fate of egg/sperm and the quality of seed produced...
August 15, 2017: BMC Genomics
https://www.readbyqxmd.com/read/28807016/endogenous-controls-of-gene-expression-in-n-methyl-n-nitrosourea-induced-t-cell-lymphoma-in-p53-deficient-mice
#3
Xi Wu, Susu Liu, Jianjun Lyu, Shuya Zhou, Yanwei Yang, Chenfei Wang, Wenda Gu, Qin Zuo, Baowen Li, Changfa Fan
BACKGROUND: Real-time polymerase chain reaction (PCR) has become an increasingly important technique for gene expression profiling because it can provide insights into complex biological and pathological processes and be used to predict disease or treatment outcomes. Although normalized data are necessary for an accurate estimation of mRNA expression levels, several pieces of evidence suggest that the expression of so-called housekeeping genes is not stable. This study aimed to validate reference genes for the normalization of real-time PCR in an N-methyl-N-nitrosourea (MNU)-induced T-cell lymphoma mouse model...
August 14, 2017: BMC Cancer
https://www.readbyqxmd.com/read/28763741/expression-stability-and-selection-of-optimal-reference-genes-for-gene-expression-normalization-in-early-life-stage-rainbow-trout-exposed-to-cadmium-and-copper
#4
Kamran Shekh, Song Tang, Som Niyogi, Markus Hecker
Gene expression analysis represents a powerful approach to characterize the specific mechanisms by which contaminants interact with organisms. One of the key considerations when conducting gene expression analyses using quantitative real-time reverse transcription-polymerase chain reaction (qPCR) is the selection of appropriate reference genes, which is often overlooked. Specifically, to reach meaningful conclusions when using relative quantification approaches, expression levels of reference genes must be highly stable and cannot vary as a function of experimental conditions...
September 2017: Aquatic Toxicology
https://www.readbyqxmd.com/read/28761164/identification-of-reference-genes-for-qrt-pcr-in-granulosa-cells-of-healthy-women-and-polycystic-ovarian-syndrome-patients
#5
Yue Lv, Shi Gang Zhao, Gang Lu, Chi Kwan Leung, Zhi Qiang Xiong, Xian Wei Su, Jin Long Ma, Wai Yee Chan, Hong Bin Liu
Comparative gene expression analysis by qRT-PCR is commonly used to detect differentially expressed genes in studies of PCOS pathology. Impaired GC function is strongly associated with PCOS pathogenesis, and a growing body of studies has been dedicated to identifying differentially expressed genes in GCs in PCOS patients and healthy women by qRT-PCR. It is necessary to validate the expression stability of the selected reference genes across the tested samples for target gene expression normalization. We examined the variability and stability of expression of the 15 commonly used reference genes in GCs from 44 PCOS patients and 45 healthy women using the GeNorm, BestKeeper, and NormFinder statistical algorithms...
July 31, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28754446/selection-of-reliable-reference-genes-for-gene-expression-studies-in-trichoderma-afroharzianum-ltr-2-under-oxalic-acid-stress
#6
Yuping Lyu, Xiaoqing Wu, He Ren, Fangyuan Zhou, Hongzi Zhou, Xinjian Zhang, Hetong Yang
An appropriate reference gene is required to get reliable results from gene expression analysis by quantitative real-time reverse transcription PCR (qRT-PCR). In order to identify stable and reliable reference genes in Trichoderma afroharzianum under oxalic acid (OA) stress, six commonly used housekeeping genes, i.e., elongation factor 1, ubiquitin, ubiquitin-conjugating enzyme, glyceraldehyde-3-phosphate dehydrogenase, α-tubulin, actin, from the effective biocontrol isolate T. afroharzianum strain LTR-2 were tested for their expression during growth in liquid culture amended with OA...
July 25, 2017: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/28746411/selection-and-validation-of-reference-genes-for-quantitative-real-time-pcr-analysis-under-different-experimental-conditions-in-the-leafminer-liriomyza-trifolii-diptera-agromyzidae
#7
Ya-Wen Chang, Jing-Yun Chen, Ming-Xing Lu, Yuan Gao, Zi-Hua Tian, Wei-Rong Gong, Wei Zhu, Yu-Zhou Du
Liriomyza trifolii is a highly-invasive leafmining insect that causes significant damage to vegetables and horticultural crops worldwide. Relatively few studies have quantified gene expression in L. trifolii using real-time quantitative PCR (RT-qPCR), which is a reliable and sensitive technique for measuring gene expression. RT-qPCR requires the selection of reference genes to normalize gene expression data and control for internal differences between samples. In this study, nine housekeeping genes from L. trifolii were selected for their suitability in normalizing gene expression using geNorm, Normfinder, BestKeeper, the ΔCt method and RefFinder...
2017: PloS One
https://www.readbyqxmd.com/read/28743297/identification-of-reference-mirnas-in-plasma-useful-for-the-study-of-oestrogen-responsive-mirnas-associated-with-acquired-protein-s-deficiency-in-pregnancy
#8
J W Tay, I James, Q W Hughes, J Y Tiao, R I Baker
BACKGROUND: Accumulating evidence indicate that circulating microRNAs (miRNAs) are useful independent non-invasive biomarkers, with unique miRNA signatures defined for various pathophysiological conditions. However, there are no established universal housekeeping miRNAs for the normalisation of miRNAs in body fluids. We have previously identified an oestrogen-responsive miRNA, miR-494, in regulating the anticoagulant, Protein S, in HuH-7 liver cells. Moreover, increased thrombotic risk associated with elevated circulating oestrogen levels is frequently observed in pregnant women and oral contraceptive users...
July 25, 2017: BMC Research Notes
https://www.readbyqxmd.com/read/28732075/comprehensive-selection-of-reference-genes-for-quantitative-rt-pcr-analysis-of-murine-extramedullary-hematopoiesis-during-development
#9
Giuliana Medrano, Peihong Guan, Amanda J Barlow-Anacker, Ankush Gosain
The purpose of this study was to perform a comprehensive evaluation and selection of reference genes for the study of extramedullary hematopoiesis during development and the early post-natal period. A total of six candidate reference genes (ACTB, GAPDH, HPRT1, PPID, TBP, TUBB3) in four organs (heart, liver, spleen, and thymus) over five perinatal time points (Embryonic days 14.5, 16.5, 18.5, Post-natal days 0, 21) were evaluated by quantitative real-time PCR. The expression stability of the candidate reference genes were analyzed using geNorm, NormFinder, Bestkeeper, Delta CT method, and RefFinder software packages...
2017: PloS One
https://www.readbyqxmd.com/read/28731643/transcriptome-based-identification-of-the-optimal-reference-cho-genes-for-normalisation-of-qpcr-data
#10
Adam J Brown, Suzanne Gibson, Diane Hatton, David C James
Real-time quantitative PCR (qPCR) is the standard method for determination of relative changes in mRNA transcript abundance. Analytical accuracy, precision, and reliability are critically dependent on the selection of internal control reference genes. In this study we have identified optimal reference genes that can be utilized universally for qPCR analysis of CHO cell mRNAs. Initially, transcriptomic datasets were analysed to identify eight endogenous genes that exhibited high expression stability across four distinct CHO cell lines sampled in different culture phases...
July 21, 2017: Biotechnology Journal
https://www.readbyqxmd.com/read/28727054/rapid-communication-mir-92a-as-a-housekeeping-gene-for-analysis-of-bovine-mastitis-related-microrna-in-milk
#11
Y C Lai, T Fujikawa, T Ando, G Kitahara, M Koiwa, C Kubota, N Miura
Our aim was to identify a suitable microRNA housekeeping gene for real-time PCR analysis of bovine mastitis-related microRNA in milk. We identified , , and as housekeeping gene candidates on the basis of previous Solexa sequencing results. Threshold cycle (CT) values for , , and did not differ between milk from control cows and milk from mastitis-affected cows. NormFinder software identified as the most stable single housekeeping gene. We evaluated the suitability of the housekeeping gene candidates by using them to assess expression levels of the inflammation-related gene ...
June 2017: Journal of Animal Science
https://www.readbyqxmd.com/read/28706523/identification-of-optimal-reference-genes-for-normalization-of-qpcr-analysis-during-pepper-fruit-development
#12
Yuan Cheng, Xin Pang, Hongjian Wan, Golam J Ahammed, Jiahong Yu, Zhuping Yao, Meiying Ruan, Qingjing Ye, Zhimiao Li, Rongqing Wang, Yuejian Yang, Guozhi Zhou
Due to its high sensitivity and reproducibility, quantitative real-time PCR (qPCR) is practiced as a useful research tool for targeted gene expression analysis. For qPCR operations, the normalization with suitable reference genes (RGs) is a crucial step that eventually determines the reliability of the obtained results. Although pepper is considered an ideal model plant for the study of non-climacteric fruit development, at present no specific RG have been developed or validated for the qPCR analyses of pepper fruit...
2017: Frontiers in Plant Science
https://www.readbyqxmd.com/read/28702046/selection-and-validation-of-appropriate-reference-genes-for-qrt-pcr-analysis-in-isatis-indigotica-fort
#13
Tao Li, Jing Wang, Miao Lu, Tianyi Zhang, Xinyun Qu, Zhezhi Wang
Due to its sensitivity and specificity, real-time quantitative PCR (qRT-PCR) is a popular technique for investigating gene expression levels in plants. Based on the Minimum Information for Publication of Real-Time Quantitative PCR Experiments (MIQE) guidelines, it is necessary to select and validate putative appropriate reference genes for qRT-PCR normalization. In the current study, three algorithms, geNorm, NormFinder, and BestKeeper, were applied to assess the expression stability of 10 candidate reference genes across five different tissues and three different abiotic stresses in Isatis indigotica Fort...
2017: Frontiers in Plant Science
https://www.readbyqxmd.com/read/28699725/evaluation-of-candidate-reference-genes-for-quantitative-expression-studies-in-asian-seabass-lates-calcarifer-during-ontogenesis-and-in-tissues-of-healthy-and-infected-fishes
#14
Anutosh Paria, Jie Dong, P P Suresh Babu, M Makesh, Aparna Chaudhari, A R Thirunavukkarasu, C S Purushothaman, K V Rajendran
Quantitative real-time PCR (qRT-PCR), used to determine the gene expression profile, is an important tool in functional genomic research, including fishes. To obtain more robust and meaningful result, the best possible normalization of the data is of utmost significance. In the present study, we have evaluated the potential of five commonly used housekeeping genes i.e., elongation factor 1-α (EF1A), β-Actin (ACTB), 18S ribosomal RNA (18S), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and β-2-Microglobulin (B2M) in normal physiological conditions, developmental stages and in response to bacterial infection in Asian seabass, Lates calcarifer (Bloch), an important food fish cultured in the Asia-Pacific region...
September 2016: Indian Journal of Experimental Biology
https://www.readbyqxmd.com/read/28695409/selection-and-validation-of-reference-genes-for-rt-qpcr-indicates-that-juice-of-sugarcane-varieties-modulate-the-expression-of-c-metabolism-genes-in-the-endophytic-diazotrophic-herbaspirillum-rubrisubalbicans-strain-hcc103
#15
Valéria Polese, Cleiton de Paula Soares, Paula Renata Alves da Silva, Jean Luiz Simões-Araújo, José Ivo Baldani, Marcia Soares Vidal
Quantitative reverse transcription PCR (RT-qPCR) is an important tool for evaluating gene expression. However, this technique requires that specific internal normalizing genes be identified for different experimental conditions. To date, no internal normalizing genes are available for validation of data analyses for Herbaspirillum rubrisubalbicans strain HCC103, an endophyte that is part of the sugarcane consortium inoculant. This work seeks to identify and evaluate suitable reference genes for gene expression studies in HCC103 grown until middle log phase in sugarcane juice obtained from four sugarcane varieties or media with three different carbon sources...
July 10, 2017: Antonie Van Leeuwenhoek
https://www.readbyqxmd.com/read/28677248/identification-of-suitable-reference-genes-during-the-formation-of-chlamydospores-in-clonostachys-rosea-67-1
#16
Jun Zhang, Zhanbin Sun, Shidong Li, Manhong Sun
Clonostachys rosea is a potential biocontrol fungus that can produce highly resistant chlamydospores under specific conditions. To investigate the genes related to chlamydospore formation, we identified reliable reference genes for quantification of gene expression in C. rosea 67-1 during sporulation. In this study, nine reference genes, actin (ACT), elongation factor 1 (EF1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), histone (HIS), RNA polymerase II CTD phosphatase Fcp1 (RPP), succinate-semialdehyde dehydrogenase (SSD), TATA-binding protein (TBP), ubiquitin (UBQ), and ubiquitin-conjugating enzyme (UCE), were selected and cloned from 67-1, and their expression stability during chlamydospore formation was determined using reverse transcription quantitative PCR and assessed using the software geNorm, NormFinder and BestKeeper...
July 5, 2017: MicrobiologyOpen
https://www.readbyqxmd.com/read/28672012/identification-and-validation-of-reference-genes-for-qrt-pcr-studies-of-the-obligate-aphid-pathogenic-fungus-pandora-neoaphidis-during-different-developmental-stages
#17
Shutao Zhang, Chun Chen, Tingna Xie, Sudan Ye
The selection of stable reference genes is a critical step for the accurate quantification of gene expression. To identify and validate the reference genes in Pandora neoaphidis-an obligate aphid pathogenic fungus-the expression of 13classical candidate reference genes were evaluated by quantitative real-time reverse transcriptase polymerase chain reaction(qPCR) at four developmental stages (conidia, conidia with germ tubes, short hyphae and elongated hyphae). Four statistical algorithms, including geNorm, NormFinder, BestKeeper and Delta Ct method were used to rank putative reference genes according to their expression stability and indicate the best reference gene or combination of reference genes for accurate normalization...
2017: PloS One
https://www.readbyqxmd.com/read/28671969/reference-gene-selection-for-the-shell-gland-of-laying-hens-in-response-to-time-points-of-eggshell-formation-and-nicarbazin
#18
Sami Samiullah, Juliet Roberts, Shu-Biao Wu
Ten reference genes were investigated for normalization of gene expression data in the shell gland of laying hens. Analyses performed with geNorm revealed that hypoxanthine phosphoribosyltransferase 1 (HPRT1) and hydroxymethylbilane synthase (HMBS) were the two most stable reference genes in response to post-oviposition time alone (POT) or with nicarbazin treatment (POT+N) of laying hens. NormFinder analyses showed that the two most stable reference genes in response to POT and POT+N were 18S ribosomal RNA (18S rRNA), ribosomal protein L4 (RPL4) and HMBS, RPL4, respectively...
2017: PloS One
https://www.readbyqxmd.com/read/28671258/selection-of-candidate-reference-genes-and-validation-for-real-time-pcr-studies-in-rice-plants-exposed-to-low-temperatures
#19
P A Auler, L C Benitez, M N do Amaral, I L Vighi, G S Rodrigues, L C da Maia, E J B Braga
Rice is a cereal that presents a great ability to adapt to different soil and climate conditions. However, as it is a tropical crop with C3 metabolism, it performs better in warm temperatures with high solar radiation. Tolerance to stress caused by low temperatures is a highly complex process that involves various metabolic pathways and cellular compartments, resulting in general or specific effects on plant growth and development. In order to observe the true effect of a particular stress on genetic expression, reference genes need to be chosen for real-time PCRs, the expression levels of which should remain stable independent of the situation imposed...
June 29, 2017: Genetics and Molecular Research: GMR
https://www.readbyqxmd.com/read/28655566/determination-of-suitable-reference-genes-for-rt-qpcr-analysis-of-murine-cytomegalovirus-in-vivo-and-in-vitro
#20
Marion Griessl, Michael Gutknecht, Charles H Cook
Reverse transcription quantitative PCR (RT-qPCR) is the most commonly used method to evaluate gene expression. Reliable qPCR results are highly dependent on accurate normalization using suitable reference genes. We investigated expression of commonly used reference genes during murine Cytomegalovirus (mCMV) infection and latency to determine those genes least perturbed by infection. Following mCMV infection in BALB/c mice, lung, salivary gland, liver, spleen and kidney were evaluated. Liver sinusoidal endothelial cells and NIH-3T3 cells were also evaluated...
June 26, 2017: Journal of Virological Methods
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