EXPERIMENT_TITLE: GSM3735449: WT-HiC-seq; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: 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.
STUDY_DESCRIPTOR_STUDY_ABSTRACT: We found that posterior HOXA-associated HOTTIP lncRNA is aberrantly activated in MLL-rearranged AMLs and required for the posterior HOXA chromatin structure and gene expression. Knock-out of HOTTIP attenuates leukemic progressions of the transplanted humanized AML mice by blocking posterior HOXA-associated AML gene expression programs while the dCas-VP160 mediated reactivation of HOTTIP restores HOXA locus chromatin structure and HOXA9-A13 gene expression in the CBS7/9 boundary depleted AML cells. Finally, transgenic expression of HOTTIP lncRNA in mouse bone marrow hematopoietic cells resulted in perturbation of the balance of HSC self-renewal and differentiation and development of AML like disease by aberrant altering HOXA associated chromatin structure and transcription program. Thus, HOTTIP lncRNA acts as oncogene to reprogram leukemic associated chromatin and gene transcription. Overall design: We have finished the RNA-SEQ, ATAC-seq, ChIP-seq, CHIRP-seq, HiC-seq and whole exome-seq to investigate the role of lncRNA HOTTIP and CTCF boundaries in hematopoiesis and leukemogenesis. LSK cells (Lin- Sca1+ Kit+) were sorted by FACS from the bone marrow of  WT and Hottip transgenic mice for RNA-seq, ATAC-seq assay. MOLM13 leukemia cells were used to perform the CHIRP-seq, RNA-seq, CHIP-seq and HiC-seq analysis.
SAMPLE_TITLE: WT-HiC-seq
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "MOLM13 leukemia cells"}, {"TAG": "cell line", "VALUE": "MOLM13"}, {"TAG": "cell types", "VALUE": "Human-derived acute myeloid leukemia cells"}, {"TAG": "tissue origin", "VALUE": "peripheral blood"}, {"TAG": "genotype/variation", "VALUE": "WT"}]
Pool_Member_@attributes_sample_title: WT-HiC-seq
RUN_SET_RUN_Pool_Member_@attributes_sample_title: WT-HiC-seq
RUN_SET_RUN_SRAFiles_SRAFile: [{"Alternatives": null, "@attributes": {"cluster": "public", "filename": "WT-HiC-seq_R1.fastq.gz", "size": "65638621770", "date": "2020-01-24 19:28:56", "md5": "b2805805d95d6a52e1b7b92be7f80894", "version": "1", "semantic_name": "fastq", "supertype": "Original", "sratoolkit": "0"}}, {"Alternatives": [null, null, null], "@attributes": {"cluster": "public", "filename": "SRR8952862.lite", "url": "https://sra-downloadb.be-md.ncbi.nlm.nih.gov/sos5/sra-pub-zq-11/SRR008/952/SRR8952862/SRR8952862.lite.1", "size": "65115851560", "date": "2022-06-10 07:06:18", "md5": "da06583fef041994f241fbb69a19f196", "version": "1", "semantic_name": "SRA Lite", "supertype": "Primary ETL", "sratoolkit": "1"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "SRR8952862", "url": "https://sra-pub-run-odp.s3.amazonaws.com/sra/SRR8952862/SRR8952862", "size": "82289256976", "date": "2019-04-25 06:14:53", "md5": "2f523b5b3e465c8eb97c65f2cfb53bc7", "version": "1", "semantic_name": "SRA Normalized", "supertype": "Primary ETL", "sratoolkit": "1"}}, {"Alternatives": null, "@attributes": {"cluster": "public", "filename": "WT-HiC-seq_R2.fastq.gz", "size": "67911972926", "date": "2020-01-25 01:45:43", "md5": "16bb3519b786a8e92d631f6634d75b08", "version": "1", "semantic_name": "fastq", "supertype": "Original", "sratoolkit": "0"}}]