Unknown,Transcriptomics,Genomics,Proteomics

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RNA-sequencing analysis of liver from mice overexpressing miR-30c

ABSTRACT: This study aims to investigate the role of microRNA-30c on hepatic and metabolic gene expression and physiology For this experiment, we used male C57BL/6 mice. At an age of 8 weeks, we started them on Western diet for one month and then injected them with either PBS or increasing dose of Scr or miR-30c mimic (2.5, 5.0, and 7.5 mg/kg) for 6 weeks. Liver from these mice were harvested and flash frozen. RNA from the livers of these mice were extracted and RNA-seq was performed.

ORGANISM(S): Mus musculus

SUBMITTER: Bailey Peck

PROVIDER: E-GEOD-70352 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:CCAT1-L is a highly expressed long noncoding RNA located in the colorectal cancer specific super enhance region about 500 kb upstream of MYC gene. Knockdown of CCAT1-L significantly down-regulated interaction frequency between CCAT1 and MYC locus and repress MYC expression, suggesting a long-range chromatin interaction between CCAT1-L and MYC locus maintained by CCAT1-L underlie the MYC regulation. To further validate this hypothesis, multiplexed 3C sequencing (3C-seq) was employed to evaluate chromatin interaction strength between CCAT1-L and MYC locus in CCAT1-L knockdown and scramble knockdown (Scr) HT29 cells. The 3C-Seq design and data analysis were performed according to Stadhouders et al, Nat Protoc. 2013, 8:509-524. A series of bait sequences accommodating different locus around CCAT1-L and MYC were selected. Through integrating with specific sample barcodes, bait-specific primer sets were designed to construct relevant 3C-seq libraries in CCAT1-L knockdown and scramble knockdown (Scr) HT29 samples. All of the 3C sample libraries from different treatment, including CCAT1-L knockdown and scramble knockdown (Scr), were then pooled together for high-throughput sequencing. Two technical 3C-seq were performed (CCAT1_myc_3C_1.txt.gz and CCAT1_myc_3C_2.txt.gz) and then combined together to get enough reads for further analysis. 3C-seq reads from different samples were divided according to sample barcodes (CCAT1-L knockdown: ATGTCA; Scr: GCCAAT) and different bait sequences, and then mapped to human reference genome to assess chromatin interaction strength between CCAT1-L and MYC locus in different treatments. In our study, one representative bait-specific sequencing data (CTTCTACTGATTGGCCCTAAACACTTTTCCAAAGCTT) was select to generate bedgraph files and upload to UCSC for visualization to show the chromatin interaction between CCAT1-L and Myc locus in CCAT1-L knockdown (CCAT1-L_knockdown_out.bedgraph) and scramble knockdown (Scr_out.bedgraph) samples.

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RNA-sequencing analysis of liver from mice overexpressing miR-30c

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