EXPERIMENT_TITLE: GSM8224471: Breast_tumor_HiC; Homo sapiens; Hi-C
EXPERIMENT_DESIGN_LIBRARY_DESCRIPTOR_LIBRARY_CONSTRUCTION_PROTOC: Breast tissue was acquired from three Tru-Cut biopsies. The tissue was sectioned into small fragments and cryopreserved in FBS (Gibco) supplemented with 10% dimethyl sulfoxide (DMSO, Sigma). Upon thawing, the tissue was enzymatically dissociated using Accumax solution (Sigma) and incubated for 30 minutes. The resulting cell suspension was filtered through a 40 µm filter, followed by processing using the Dead Cell Removal Kit (Miltenyi Biotec #130090101) through two rounds to ensure sufficient viability. Cells were cross-linked with 2% formaldehyde. Then cell nuclei were obtained with a lysis buffer (10 mM Tris-HCl pH 8, 0.2% IGEPAL CA-630, 10 mM NaCl, 1X cOmpleteR protease inhibitors cocktail). For genome digestion, 125 U of DpnII (New England Biolabs) were added. The ligation spet was performed using 50 U Weiss of T4 DNA Ligase enzyme (Thermo Scientific, 5 U Weiss/μl), and the crosslink was reversed. DNA was purified using a Phenol-Chloroform-Isoamyl alcohol mix (25:24:1) and shared with a Covaris Sonicator. Hi-C libraries were generated using the TruSeq Illumina system.
STUDY_DESCRIPTOR_STUDY_ABSTRACT: Molecular and cellular characterization of tumors has become essential due to the complex and heterogeneous nature of cancer disease. In recent decades, many bioinformatic tools and experimental techniques have been developed to achieve personalized characterization of tumors. However, sample handling continues to be a major challenge as limitations such as prior treatments before sample acquisition, the amount of tissue obtained, transportation, or the inability to process fresh samples pose a challenge for experimental strategies that require viable cell suspensions. Here, we present an optimized protocol that allows recovery of highly viable cell suspensions from breast cancer biopsies. Using these cell suspensions we successfully characterized genome architecture through Hi-C. Also, we evaluated single-cell gene expression and the tumor cellular microenvironment through single cell RNAseq. Both technologies are key in the detailed and personalized molecular characterization of tumor samples. The protocol described here opens the possibility to obtain viable cell suspensions from biopsies in a simple and efficient manner Overall design: The cell suspension was derived from a Tru-Cut biopsy sample obtained from a Mexican female patient diagnosed with untreated triple-negative breast cancer.
SAMPLE_TITLE: Breast_tumor_HiC
SAMPLE_SAMPLE_ATTRIBUTES_SAMPLE_ATTRIBUTE: [{"TAG": "source_name", "VALUE": "Breast cancer primary tumor"}, {"TAG": "tissue", "VALUE": "Breast cancer primary tumor"}, {"TAG": "cell type", "VALUE": "Triple negative breast cancer"}, {"TAG": "geo_loc_name", "VALUE": "missing"}, {"TAG": "collection_date", "VALUE": "missing"}]
Pool_Member_@attributes_sample_title: Breast_tumor_HiC
RUN_SET_RUN_Pool_Member_@attributes_sample_title: Breast_tumor_HiC
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