Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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MiRNA Microarray Profiling with Three Different Total RNA Prep Methods

ABSTRACT: We performed miRNA microarray profiling on samples prepared from two different cell lines by three widely-used total RNA sample prep methods. miRNA microarrays were manufactured by Agilent Technologies (Santa Clara, CA), and contain 20-40 features targeting each of 470 human miRNAs (Agilent design ID 016436). Sequences of the 470 miRNAs were obtained from the Sanger miRBase, release 9.1. Labeling and hybridization of total RNA samples were performed according to the manufacturer’s protocol. 100ng total RNA was used as input into the labeling reaction, and the entire reaction was hybridized to the array for 20 hours at 55°C. For total RNA preps, frozen cell pellets were resuspended in phosphate buffered saline and divided into equal aliquots of 5x106 (HeLa) or 1x107 (ZR-75-1) cells and refrozen. Individual aliquots were subsequently thawed just before use.

ORGANISM(S): Homo sapiens

SUBMITTER: Robert Ach

PROVIDER: E-GEOD-11840 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Measuring microRNAs: comparisons of microarray and quantitative PCR measurements, and of different total RNA prep methods.

Ach Robert A RA Wang Hui H Curry Bo B

BMC biotechnology 20080911

<h4>Background</h4>Determining the expression levels of microRNAs (miRNAs) is of great interest to researchers in many areas of biology, given the significant roles these molecules play in cellular regulation. Two common methods for measuring miRNAs in a total RNA sample are microarrays and quantitative RT-PCR (qPCR). To understand the results of studies that use these two different techniques to measure miRNAs, it is important to understand how well the results of these two analysis methods cor ...[more]

PMID: 18783629

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Project description:On going efforts are directed at understanding the mutualism between the gut microbiota and the host in breast-fed versus formula-fed infants. Due to the lack of tissue biopsies, no investigators have performed a global transcriptional (gene expression) analysis of the developing human intestine in healthy infants. As a result, the crosstalk between the microbiome and the host transcriptome in the developing mucosal-commensal environment has not been determined. In this study, we examined the host intestinal mRNA gene expression and microbial DNA profiles in full term 3 month-old infants exclusively formula fed (FF) (n=6) or breast fed (BF) (n=6) from birth to 3 months. Host mRNA microarray measurements were performed using isolated intact sloughed epithelial cells in stool samples collected at 3 months. Microbial composition from the same stool samples was assessed by metagenomic pyrosequencing. Both the host mRNA expression and bacterial microbiome phylogenetic profiles provided strong feature sets that clearly classified the two groups of babies (FF and BF). To determine the relationship between host epithelial cell gene expression and the bacterial colony profiles, the host transcriptome and functionally profiled microbiome data were analyzed in a multivariate manner. From a functional perspective, analysis of the gut microbiota's metagenome revealed that characteristics associated with virulence differed between the FF and BF babies. Using canonical correlation analysis, evidence of multivariate structure relating eleven host immunity / mucosal defense-related genes and microbiome virulence characteristics was observed. These results, for the first time, provide insight into the integrated responses of the host and microbiome to dietary substrates in the early neonatal period. Our data suggest that systems biology and computational modeling approaches that integrate “-omic” information from the host and the microbiome can identify important mechanistic pathways of intestinal development affecting the gut microbiome in the first few months of life. KEYWORDS: infant, breast-feeding, infant formula, exfoliated cells, transcriptome, metagenome, multivariate analysis, canonical correlation analysis 12 samples, 2 groups

Dataset Information

MiRNA Microarray Profiling with Three Different Total RNA Prep Methods

Publications

Measuring microRNAs: comparisons of microarray and quantitative PCR measurements, and of different total RNA prep methods.

Similar Datasets

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