MRNA-seq of DND-41 T-cell acute lymphoblastic leukemia cell line upon inhibition of hsa-miR-363-3p
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ABSTRACT: The aim of this study was to investigate the effect of hsa-miR-363-3p on global gene expression in T-cell acute lymphoblastic leukemia (T-ALL). For this purpose we selected DND-41 T-ALL cell line characterized by high expression of this miRNA. The cells were transduced either with miRZip pGreenPuro scrambled vector (System Biosciences) or miRZip anti-hsa-miR-363-3p vector, each variant in three biological replicates. All 6 libraries were sequenced on Illumina NovaSeq6000 platform, in 2x150 mode, with a coverage of 60M paired reads/sample.
Project description:The aim of this study was to investigate the effect of hsa-miR-143-3p on global gene expression in T-cell acute lymphoblastic leukemia (T-ALL) and to unravel its direct target genes. For this purpose we selected Jurkat T-ALL cell line characterized by low expression of this miRNA. The cells were transduced either with pCDH-CMV-MCS-EF1_GFP_puro empty vector (System Biosciences) or pCDH-CMV-MCS-EF1_GFP pre-mir-143 vector, each variant in three biological replicates. Total RNA was isolated from each samples. Additionally, RISC complexes containing miRNAs bound to their target mRNAs were isolated by AGO2 immunoprecipitation and the obtained fraction of RISC-bound RNA was isolated from each sample. All 12 libraries (6 total RNA samples and 6 AGO2-RIP samples) were sequenced on Illumina NovaSeq6000 platform, in 2x150 mode, with a coverage of 60M paired reads/sample.
Project description:The aim of this study was to investigate the transcriptome of 54 primary samples of pediatric patients diagnosed with T-cell acute lymphoblastic leukemia. Samples were collected before treatment. Sequencing libraries were obtained with Illumina TruSeq Stranded mRNA protocol, with modified conditions of RNA fragmentation (90°C for 2 minutes). All libraries were sequenced on Illumina NovaSeq6000 platform, in 2x150PE mode (paired end sequencing with 150 nt reads), with a coverage of 150M reads/sample.
Project description:The aim of this study was to compare two different library preparation protocols: Illumina TruSeq Stranded mRNA (referred to as PA) and TruSeq Stranded Total RNA with Ribo-Zero Human/Mouse/Rat (RD) each with two different variants of the fragmentation step: standard fragmentation conditions (94°C for 8 minutes; further described as P1) and modified conditions (90°C for 2 minutes; further described as P2). The modification was expected to produce longer fragments. Each of the 5 tested T-ALL samples was sequenced using a combination of 4 different protocols: PA_P1, RD_P1, PA_P2, RD_P2 (for PA_P2 we performed two technical replicates). All 25 libraries were sequenced on Illumina NovaSeq6000 platform, in 2x150 mode, with a coverage of 150M reads/sample. High number of reads obtained for each of the samples allowed us to carry out a comprehensive comparison using various analysis methods aimed at identifying differentially expressed genes, alternative splicing events, gene fusions, somatic mutations and indels.
Project description:The aim of this study was to analyze gene expression of primary airway epithelial cells during their differentiation in air-liquid interface ALI culture. Cells from one healthy donor were collected at different days of ALI culture (0,6,9,14 and 21). All samples were sequenced on Illumina NovaSeq6000 platform, in 2x150 mode, with a coverage of 100M reads/sample. Obtained data was later correlated with expression of 14 genes known to be related to airway epithelium differentiation obtained using qRT-PCR technique (TaqMan low density array, TLDA cards).
Project description:The requirement for primordial germ cells (PGCs) during sexual differentiation is variable among vertebrates. It has been shown that in zebrafish complete loss of PGCs in the embryos causes exclusive male development. Further, transplantation of a single PGC into a germline deficient zebrafish embryos generates male exclusively, suggesting PGC number might be important for the ovarian fate.To explore how PGC number might regulate sexual development in zebrafish, we experimentally manipulated its number by injecting dnd MO into embryos to generate fish containing a spectrum of PGC number. The experiment was designed to compare transcriptomes between the developing trunk regions of wild type and dnd morphants at different developmental stages. The dnd MO was microinjected into one cell stage embryos to generate zebrafish with a range of PGC numbers. Transcriptomes of developing trunk regions of wild type and dnd morphants at 14 dpf and 22 dpf were analyzed.
Project description:Background: The identification of new high sensitivity and specificity markers for HCC are essential. We aimed to identify serum microRNAs for diagnosing hepatitis B virus (HBV) â??related HCC. Methods: Serum microRNA expression was investigated with four cohorts including 667 participants (261 HCC patients ,233 cirrhosi patients and 173 healthy controls), recruited between August 2010 and June 2013. First, An initial screening of miRNA expression by Illumina sequencing was performed using serum samples pooled from HCC patients and controls,respectively. Quantitative reverse-transcriptase polymerase chain reaction assay was then applied to evaluate the expression of selected microRNAs. A logistic regression model was constructed using a training cohort (n=357) and then validated using a cohort(n=241). The area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. Results: , We identified 8 miRNAs(hsa-miR-206, hsa-miR-141-3p, hsa-miR-433-3p, hsa-miR-1228-5p, hsa-miR-199a-5p, hsa-miR-122-5p, hsa-miR-192-5p and hsa-miR-26a-5p.) formed a miRNA panel that provided a high diagnostic accuracy of HCC (AUC=0.887 and 0.879 for training and validation data set, respectively). The microRNA panel can also differentiate HCC from healthy (AUC =0.894) and cirrhosis (AUC = 0.892), respectively. Conclusions:We found a serum microRNAs panel that has considerable clinical value in diagnosing HCC. 9 serum samples pooled from 3 healthy control donors and 3 HCC patients, 3 cirrhosi patients treated at The First Affiliated Hospital of Soochow University were subjected to Illumina HiSeq 2000 deep sequencing to identify the miRNAs that were significantly differentially expressed.
Project description:Microglia were derived from iPSCs and treated with mimics and inhibitors of the miRNAs hsa-miR-150-5p, hsa-miR-193a-3p and hsa-miR-19b-3p. RNA-sequencing was then performed to examine the effects of up- and down-regulation of the respective miRNAs.
Project description:iPSC-derived neurons were treated with mimics and inhibitors of the miRNAs miR-150-5p, hsa-mir-193a-3p and hsa-miR-19b-3p.
RNA-sequencing was then performed to examine the effects of miRNA up-regulation and inhibition.
Project description:In our previous study, hsa-let-7d-3p, hsa-let-7e-5p,hsa-miR-146a-5p,hsa-miR-130a-3p, hsa-miR-151a-3p,were significantly upregulated in the plasma of atopic patients. To study the each function of let-7d-3p, let-7e-5p,miR-146a-5p,miR-130a-3p, miR-151a-3p which are significantly upregulated in the plasma of atopic patients, we performed mimic-transfected THP-1 cells, a mononuclear cell line, and performed comprehensive genetic analysis.