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Sample GSM3972132 Query DataSets for GSM3972132
Status Public on Jul 23, 2020
Title T-M4
Sample type SRA
Source name Whole Tumor-Burdened Lung
Organism Mus musculus
Characteristics strain: mixed; C57/BL/6J; 129/SV
tumor stage: late-stage tumor
Treatment protocol Mice with tumors were induced at age 6-10 weeks. Mice were provided Adeno-SPC-Cre virus intratracheally resulting in recombination of Kras G12D, p53, and tdTomato in alveolar type II cells.
Growth protocol 3T3 cells were grown in DMEM + 10% FBS + 1% pen/strep; GM12878 cells were grown in RPMI + 10% FBS + 1% pen/strep
Extracted molecule genomic DNA
Extraction protocol Lungs were dissociated with fine scissors and then proteolytic digestion was performed using the Lung Dissociation kit (Miltenyi Biotech) following the manufacturer’s instructions. Dissociated cells were then incubated at 37 degrees C for 20 minutes with rotation, then filtered using a 100-μm strainer. Red blood cells were lysed using ACK buffer (Thermo Scientific) and stained with APC-conjugated CD45 (BD, 559864), CD11b (eBioscience 17-0112-82), CD31 (Biolegend, 102510), Ter119 (BD, 557909), and DAPI (Sigma-Aldrich). Fluorescence-activated cell sorting (FACS) of immuno-stained primary cells was performed using a FACSAria sorter (BD) to isolate tdTomato+; DAPI-; APC- tumor cells for sciATAC-seq. Cells were then processed for sciATAC-Seq profiling following 0.1% fixation.
The PCR reaction was carried out at the following conditions: 72 °C for 5 minutes (extension), 98 °C for 5 min, and then thermocycling at 98 °C for 10 s, 70 °C for 30 s and 72 °C for 1 min. After thermocycling for 5 cycles, we took 5 µl sample from a few randomly selected wells and added 10 µl of PCR cocktail with 0.6x SYBRgreen. The 15 µl reactions were amplified to saturation to determine the number of cycles required for the remaining samples on the plate. Libraries were amplified for 13-14 cycles in total. Libraries were pooled and purified using Qiagen MinElute PCR purification column. The libraries were quantified using KAPA library quantification kit. Libraries were sequenced on the Next-seq platform (Illumina) using a 150-cycle kit (Read 1: 47 cycles, Index 1: 36 cycles, Index 2: 36 cycles, Read 2: 47 cycles).
Library strategy ATAC-seq
Library source genomic
Library selection other
Instrument model Illumina NextSeq 500
Description sciATAC tumor-burdened lung
Data processing Supplementary_files_format_and_content: Excel spreadsheet containing cell and sample-specific barcodes to demultiplex pooled sequence read files in .txt format. Columns include cell number, tissue source,lLouvain cluster, index1, and index2.
Submission date Jul 24, 2019
Last update date Jul 23, 2020
Contact name Lindsay M LaFave
Organization name MIT
Department Biology
Lab Dr. Tyler Jacks
Street address 77 Massachusetts Avenue; 76-453
City Cambridge
State/province MA
ZIP/Postal code 02139
Country USA
Platform ID GPL19057
Series (2)
GSE134812 Single-cell epigenomics identifies a premetastatic regulatory program in lung adenocarcinoma [sciATAC-seq]
GSE145194 Single-cell epigenomics identifies a premetastatic regulatory program in lung adenocarcinoma
BioSample SAMN12358332
SRA SRX6585779

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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