In this study, we present a comprehensive evaluation of four RNA-Seq library preparation methods. We used three standard input protocols, the Illumina TruSeq Stranded Total RNA and TruSeq Stranded mRNA kits, and a modified NuGEN Ovation v2 kit; and an ultra-low-input RNA protocol, the TaKaRa SMARTer Ultra Low RNA Kit v3. Our evaluation of these kits included quality control measures such as overall reproducibility, 5' and 3' end-bias, and the identification of DEGs, lncRNAs, and alternatively spliced transcripts. Overall, we found that the two Illumina kits were most similar in terms of recovering DEGs, and the Illumina, modified NuGEN, and TaKaRa kits allowed identification of a similar set of DEGs. However, we also discovered that the Illumina, NuGEN and TaKaRa kits each enriched for different sets of genes. Overall design: Two mESC cell lines (biological replicates) from Zbtb24 knockout (1lox/1lox) clones are compared with two wild-type (2lox/+) clones (biological replicates) using the TaKaRa SMARTer Ultra Low RNA protocol directly on cells with no RNA preparation step. Total RNA from 100 mESCs cells and 1000 mESCs cells or approximately 1 and 10 ng RNA were used respectively.
Systematic evaluation of RNA-Seq preparation protocol performance.
Specimen part, Cell line, Subject
View SamplesIn this study, we present a comprehensive evaluation of four RNA-Seq library preparation methods. We used three standard input protocols, the Illumina TruSeq Stranded Total RNA and TruSeq Stranded mRNA kits, and a modified NuGEN Ovation v2 kit; and an ultra-low-input RNA protocol, the TaKaRa SMARTer Ultra Low RNA Kit v3. Our evaluation of these kits included quality control measures such as overall reproducibility, 5' and 3' end-bias, and the identification of DEGs, lncRNAs, and alternatively spliced transcripts. Overall, we found that the two Illumina kits were most similar in terms of recovering DEGs, and the Illumina, modified NuGEN, and TaKaRa kits allowed identification of a similar set of DEGs. However, we also discovered that the Illumina, NuGEN and TaKaRa kits each enriched for different sets of genes. Overall design: Three mESC cell lines (biological replicates) from Zbtb24 knockout (1lox/1lox) clones are compared with three wild-type (2lox/+) clones (biological replicates) using the TruSeq mRNA protocol.
Systematic evaluation of RNA-Seq preparation protocol performance.
Specimen part, Cell line, Subject
View SamplesAnopheles gambiae,the primary African malarial mosquito, exhibits numerous behaviors that are under diel and circadian control, including locomotor activity, swarming, mating, host seeking, eclosion, egg laying and sugar feeding. However, little has been performed to elucidate the molecular basis for these daily rhythms. To study how gene expression is globally regulated by diel and circadian mechanisms, we have undertaken a DNA microarray analysis ofA. gambiaehead and bodies under 12:12 light:dark cycle (LD) and constant dark (DD, free-running) conditions. Zeitgeber Time (ZT) with ZT12 defined as time of lights OFF under the light:dark cycle, and ZT0 defined as end of the dawn transition. Circadian Time (CT) with CT0 defined as subjective dawn, inferred from ZT0 of the previous light:dark cycle.
Genome-wide profiling of diel and circadian gene expression in the malaria vector Anopheles gambiae.
Sex, Specimen part
View SamplesAging within the human hematopoietic system associates with increased incidence of anemia and myeloid neoplasms, decreased bone marrow (BM) cellularity and reduced adaptive immune responses. Similar phenotypes have been observed in mice and shown, at least in part, to involve hematopoietic stem cells (HSCs). However, evidence supporting such an association within human hematopoiesis is still sparse and prompted us to detail characteristics of human hematopoietic stem and progenitor cells throughout ontogeny.
Human and Murine Hematopoietic Stem Cell Aging Is Associated with Functional Impairments and Intrinsic Megakaryocytic/Erythroid Bias.
Specimen part
View SamplesDifferent fibroblast cells (eight in total) with different inhibitory capacity were analyzed and compared for their gene expression profile by whole genome microarray.
Confrontation of fibroblasts with cancer cells in vitro: gene network analysis of transcriptome changes and differential capacity to inhibit tumor growth.
Cell line, Treatment
View SamplesWe stratified colorectal tumor samples using a new unsupervised, iterative method based on non-negative matrix factorization (NMF). The resulting five subtypes exhibited activation of specific signaling pathways, and significant differences in microsatellite status and tumor location. We could also align three CRC cell lines panels to these subtypes.
Subtypes of primary colorectal tumors correlate with response to targeted treatment in colorectal cell lines.
Sex, Race
View SamplesNitrogen and light are two major regulators of plant metabolism and development. While genes involved in the control of each of these signals have begun to be identified, regulators that integrate gene responses to nitrogen and light signals have yet to be determined.
Modeling the global effect of the basic-leucine zipper transcription factor 1 (bZIP1) on nitrogen and light regulation in Arabidopsis.
Specimen part
View SamplesAndrogen receptor (AR) is a hormone-activated transcription factor that plays important roles in prostate development, function, as well as malignant transformation. The downstream pathways of AR, however, are incompletely understood. AR has been primarily known as a transcriptional activator inducing prostate-specific gene expression. Through integrative analysis of genome-wide AR occupancy and androgen-regulated gene expression, here we report AR as a globally acting transcriptional repressor. This repression is mediated by androgen responsive elements (ARE) and dictated by Polycomb group protein EZH2 and repressive chromatin remodeling. In embryonic stem cells, AR-repressed genes are occupied by EZH2 and harbor bivalent H3K4me3 and H3K27me3 modifications that are characteristic of differentiation regulators, the silencing of which maintains the undifferentiated state. Concordantly, these genes are silenced in castration-resistant prostate cancer rendering a stem cell-like lack of differentiation and tumor progression. Collectively, our data reveal an unexpected role of AR as a transcriptional repressor inhibiting non-prostatic differentiation and, upon excessive signaling, resulting in cancerous de-differentiation. It provides an innovative mechanism for castration resistance and highlights novel therapeutic strategies to treat advanced prostate cancer.
Cooperation between Polycomb and androgen receptor during oncogenic transformation.
Cell line, Treatment
View SamplesHsa-mir-365-2 is one of the two precursors that give rise to miR-365. We discovered that miR-365 directly regulates a lung cancer and developmental gene termed thyroid transcription factor 1 (TTF-1 or NKX2-1).
MiR-365 regulates lung cancer and developmental gene thyroid transcription factor 1.
Cell line
View SamplesTo identify transcriptional targets altered in the embryonic heart after exposure to TCE, and possible protective effects of folate, we used DNA microarray technology to profile gene expression in embryonic mouse hearts with maternal TCE exposure and dietary changes in maternal folate. Results: Exposure to low doses of TCE (10ppb) caused extensive alterations in transcripts encoding proteins involved in transport, ion channel, transcription, differentiation, cytoskeleton, cell cycle and apoptosis. Exogenous folate did not offset the effects of TCE exposure on normal gene expression and both high and low levels of folate produced additional significant changes in gene expression. Conclusions: A mechanism where TCE induces a folate deficiency does not explain altered gene expression patterns in the embryonic mouse heart. The data further suggest that use of folate supplementation, in the presence of this toxin, may be detrimental and non-protective of the developing embryo.
Gene expression profiling in the fetal cardiac tissue after folate and low-dose trichloroethylene exposure.
Specimen part
View Samples