Project description:Gene expression is balanced by transcription and mRNA degradation. Shortening of polyadenosine (poly(A)) tails (deadenylation) is the initial step in the decay of most mRNAs. However, the mechanism by which deadenylation controls mRNA expression is not fully understood. This experiment aimed to determine changes in transcriptional activity in livers of control and Cnot1 tamoxifen-inducible liver-specific knockout mice. Control and Cnot1 tamoxifen-inducible liver-specific knockout mice placed on tamoxifen-containing food at 6 weeks of age for 2 weeks. Lysates from livers were subjected to ChIP assay with Histone H3 (tri methyl K4) (H3K4me3) antibody (ab8580; abcam) using SimpleChIP Enzyatic Chromatin IP Kit (#9003; Cell Signaling Technology). Libraries for DNA sequence were prepared from DNA isolated by ChIP assays with a KAPA Hyper Prep Kit (Illumina). 109 base-pair pair-end read DNA-seq was performed with Hiseq PE Rapid Cluster Kit v2-HS and Hiseq Rapid SBS Kit v2-HS (200 Cycle) on Illumina Hiseq2500.
Project description:An acRIP experiment was performed in KMS28-PE WT and KMS28-PE NAT10-OE cells. We compared the peak summit of KMS28-PE WT and KMS28-PE NAT10 OE cells to find the differences between KMS28-PE WT and KMS28-PE NAT10 OE cells.
Project description:Microspore embryogenesis is an in vitro system in which haploid microspores (precursor celsl of pollen grains) are reprogrammed towards embryogenesis by the application of a heat stress (HS) treatment. This process is of great interest in plant breeding as it enables accelerated production of double-haploid plants. The induction of in vitro microspore reprogramming comprises a complex chain of events that still remain unknown. The process can be induced at the responsive developmental stage of vacuolated microspore. In order to evaluate the transcriptional changes acompaying microspore embryogenesis induction in Brassica napus, an expression profiling through high throughput RNA sequencing, and a pipeline for bioinformatic analysis was performed. The analyses have been implemented for two samples: (1) vacuolated microspores (VM), before stress treatment, initial stage of the microspore culture, and (2) stress-treated microspores, 4 days after the application of the HS, when proembryos (PE) are formed; PE correspond to the first morphological sign of embryogenesis induction. The results obtained would provide valuable information about this complex process.
Project description:Human ovarian adenocarcinoma SKOV3 cells were exposed to BPA (10 or 100 nM) or 0.1% DMSO for 24 h,and then total RNA was extracted from cells using Trizol reagent. Sequencing libraries were generated using NEBNext UltraTM RNA Library Prep Kit for Illumina (NEB) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. Then, the index-coded samples were clustered on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Hisreq 4000 platform with 150 bp paired-end reads.
Project description:Gene expression is balanced by transcription and mRNA degradation. Shortening of polyadenosine (poly(A)) tails (deadenylation) is the initial step in the decay of most mRNAs. However, the mechanism by which deadenylation controls mRNA expression is not fully understood. This experiment aimed to determine changes in gene expression and mRNA stability in livers of control and Cnot1 tamoxifen-inducible liver-specific knockout mice. For comprehensive mRNA half-life profiling, we injected control and Cnot1 tamoxifen-inducible liver-specific knockout mice, which were placed on a tamoxifen-containing diet at 6 weeks of age for 2 weeks, intraperitoneally with 2 mg actinomycin D per g body weight for 4 and 8 hr. 1 g of total RNA was used for RNA-seq library preparation with Illumina TruSeq Stranded mRNA LT Sample Prep Kit. 109 base-pair pair-end read RNA-seq was performed with Hiseq PE Rapid Cluster Kit v2-HS and Hiseq Rapid SBS Kit v2-HS (200 Cycle) on Illumina Hiseq2500.
Project description:Heart fibroblasts from wildtype mice and Siah2-/- knockout mice were isolated and cultured. The cells were either left untreated or incubated for 6 hs under hypoxic conditions. One experiment consists of wildtype cells (normoxia/hypoxia) and Siah2 knockout cells (normoxia/hypoxia) = 4 samples. To allow statistical analysis of the data set the experiment was repeated once under identical conditions.
Project description:Purpose: The goals of this study are to compare genes change between FBXW7f/f and LysM-cre FBXW7f/f BMDM after LPS treatment Methods: BMDM mRNA from 6-8weeks FBXW7f/f (WT) and LysM-cre FBXW7f/f (KO) mice were generated by transcription sequencing, using Illumina. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform and 125 bp/150 bp paired-end reads were generated.
Project description:The risk of multiple placental thrombosis is increased in preeclampsia (PE) and vascular endothelium plays vital roles in coagulation such as secreting ITGA2B, vWF and TF. However, the pathological mechanism is unclear. Placental blood vessels were collected from PE and normal pregnancies. The protein level of ITGA2B was up-regulated in PE compared to the control, indicating enhanced coagulation function in the placenta of PE. Higher cholesterol levels and lower ALKBH1 expression were found in placental blood vessels of PE while the whole DNA N6-methyldeoxyadenosine levels (6mA) were up-regulated. However, DNA 6mA level of ITGA2B promoter was decreased. The Cholesterol overload experiment in human umbilical vein endothelial cells (HUVECs) showed that cholesterol regulated the protein expressions of ALKBH1 and ITGA2B, which was related to cholesterol concentration. ITGA2B protein expression was increased after ALKBH1 knockout. These data provide important information on cholesterol levels, ALKBH1 and ITGA2B expressions in coagulation function in placental blood vessels, which may be helpful for further investigations of potential targets and early prevention of placental thrombosis in PE.