|
| Status |
Public on Apr 29, 2024 |
| Title |
HTGTS_PC-9.pMSCV_after rep1 |
| Sample type |
SRA |
| |
|
| Source name |
HTGTS_PC-9.pMSCV_after
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: PC-9
cell type: Non-small cell lung cancer
genotype/variation: pMSCV
treatment: osimertinib
|
| Treatment protocol |
100 nM osimertinib, every other day
Cells were treated with 100 nM of osimertinib every other day.
|
| Growth protocol |
RPMI 1640 + 2mM Glutamine + 10% Fetal Bovine Serum (FBS)
PC-9 cells are grown in RPMI supplemented with 2 mM Glutamine and 10% FBS.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Rapid lysis buffer + 10ug/ml Proteinase K, at 56°C, overnight
Gennomic DNAs were extracte with Rapid lysis buffer containing 20 ug/ml of proteinase K overnight at 56°C.
Library was prepaired with the HTGTS methods previously described (Chiarle at al., Cell, 2011_PMID:21962511; Genome-wide Translocation Sequencing Reveals Mechanisms of Chromosome Breaks and Rearrangements in B Cells)
We induced DNA DSBs in intron 19 of ALK and we used linker forward primer and specific target reverse primers to capture functional fusions.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina MiSeq |
| |
|
| Data processing |
HTGTS junction reads for each sample were recognized by designed barcode, primer and bait portion as previously described (Mara Compagno et al., Nature 2017). The barcode, primer and bait portion of the remained sequences were masked for alignment analysis to determine translocation junctions. Briefly, we aligned sequences to the human genome (GRCh38/hg38) using BLAT, and then filtered artificial junctions by removing PCR repeats (reads with same junction position in alignment to the reference genome and a start position in the read less than 3 bp apart), invalid alignments (including alignment scores < 30, reads with multiple alignments having a score difference < 4 and alignments having 10-nucleotide gaps) and ligation artifacts (for example, random HaeIII restriction sites ligated to bait breaksite). Translocation junction position was determined based on the genomic position of the 5’ end of the aligned read.
Genome_build: hg38
Supplementary_files_format_and_content: HTGTS junctions; BED files contain all obtained translocation junctions information including chromosome,corrdinate,read name,align score and strand.
|
| |
|
| Submission date |
Feb 20, 2021 |
| Last update date |
Apr 29, 2024 |
| Contact name |
Roberto Chiarle |
| E-mail(s) |
Roberto.Chiarle@childrens.harvard.edu
|
| Organization name |
Children’s Hospital Boston and Harvard Medical School
|
| Department |
Department of Pathology
|
| Lab |
Roberto Chiarle
|
| Street address |
300 Longwood Avenue
|
| City |
Boston |
| State/province |
Massachusetts |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL15520 |
| Series (1) |
| GSE167155 |
Genome-wide identification of oncogenic fusions in lung cancer by functional translocation sequencing |
|
| Relations |
| BioSample |
SAMN18016429 |
| SRA |
SRX10142291 |
