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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Dec 03, 2019 |
| Title |
Hottip-Tg-ST-HSC-RNA-seq-2 |
| Sample type |
SRA |
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| Source name |
Lin- Sca1+ Kit+ cd135- cd34+ Bone marrow cells
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| Organism |
Mus musculus |
| Characteristics |
cell types: Mouse-derived Bone marrow cells
tissue origin: Bone marrow
cell subtype: Lin- Sca1+ Kit+ cd135- cd34+
genotype/variation: Hottip-Tg
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| Treatment protocol |
Immediately extract RNA from LSK, HSC cells from transgenic mice and MOLM13 AML cells without any treatment.
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| Growth protocol |
Total bone marrow cells were isolated from WT and Hottip-Tg mice, and sorted with Lin-, Sca1+ and Kit+ markers. Human MOLM13 leukemia cells were cultured at RMPI1640 and 10% FBS media.
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| Extracted molecule |
total RNA |
| Extraction protocol |
RNA samples extracted with Trizol and treated with DNase I. ChIP Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with H3K4me3, H3K27me3, H3K79me2 and MLL antibody (antibody: Anti-H3K4me3, Millipore, Cat No. 04-745; Anti-H3K27me3, Millipore, Cat No. 07-449; Anti-H3K79me2, Abcam, Cat No. ab3594.; Anti-MLL, Millipore, #05-765). ATAC-seq samples derived according to Nextera Tn5 Transposase kits. CHIRP-seq library constructs with the Illumina TruSeq CHIP DNA library kit. HiC-DNA was prepared with Arima-HiC kit (Arima, #A410030).
RNA-seq libraries were prepared according to TruSeq Stranded mRNA Library Prep (#20020594). ChIP-seq and CHIRP-seq libraries were prepared according to Illumina's instructions accompanying the TruSeq ChIP Library Preparation Kit (#IP-202-1012). ATAC-seq libraries were prepared according to Nextera DNA Library Prep Kit (#FC-121-1030). 4C-seq libraries were prepared according to TruSeq ChIP Library Preparation Kit (#IP-202-1012). HiC library was prepared according to Arima-HiC Kit (Catlog: A410030). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Standard Illumina software base-calling and quality-control filtering was applied Sequences (Paired end).
Paired end RNA-seq reads from mice were aligned to the mm9 genome assembly using Tophat, and ATAC-seq, ChIP-seq, HiC-seq and CHIRP-seq reads from human cells were aligned to the hg19 genome assembly using BOWTIE2 default parameters.
Peak calling was performed using MACS algorithm (version 2.1.1).
Genome coverage tracks were created using deepTools version 3.0.
Chromatin structure HiC-seq was calculated and displayed with HOMER, Juicer and Juicebox software.
Genome_build: mm9, hg19
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for RNA-seq Samples, and peaks annotation for ATAC-seq, ChIP-seq and CHIRP-seq.
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| Submission date |
Apr 24, 2019 |
| Last update date |
Dec 03, 2019 |
| Contact name |
Suming Huang |
| E-mail(s) |
huanglabseq@hotmail.com
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| Organization name |
Penn State University
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| Department |
Pediatrics
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| Street address |
500 University Dr.
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| City |
Hershey |
| State/province |
PA |
| ZIP/Postal code |
17033 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (1) |
| GSE114981 |
Activation of HOTTIP lncRNA perturbs HSC function leading to AML like disease |
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| Relations |
| BioSample |
SAMN11492944 |
| SRA |
SRX5732692 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3735460_Hottip-Tg-ST-HSC-RNA-seq-2_FPKM.txt.gz |
179.4 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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