概要: 10X Genomics single cell RNAseq of MCF7 cells
Human cancer cell lines are the workhorse of cancer research. While cell lines are known to evolve in culture, the extent of the resultant genetic and transcriptional heterogeneity and its functional consequences remain understudied. Here, genomic analyses of 106 cell lines grown in two laboratories revealed extensive clonal diversity. Follow-up comprehensive genomic characterization of 27 strains of the common breast cancer cell line MCF7 uncovered rapid genetic diversification. Similar results were obtained with multiple strains of 13 additional cell lines. Importantly, genetic changes were associated with differential activation of gene expression programs and marked differences in cell morphology and proliferation. Barcoding experiments showed that cell line evolution occurs as a result of positive clonal selection that is highly sensitive to culture conditions. Analyses of single cell-derived clones showed that ongoing instability quickly translates into cell line heterogeneity. Testing of the 27 MCF7 strains against 321 anti-cancer compounds uncovered strikingly disparate drug response: at least 75% of compounds that strongly inhibited some strains were completely inactive in others. This study documents the extent, origin and consequence of genetic variation within cell lines, and provides a framework for researchers to measure such variation in efforts to support maximally reproducible cancer research.
项目整体设计: Single cell clones were derived from MCF7 cells (strain L) and cultured.
MCF7 cells were cultured in RPMI-1640 (Life Technologies), with 10% Fetal Bovine Serum (Sigma-Aldrich) and 1% Penicillin-Streptomycin-Glutamine (Life Technologies).
处理方案:
Single cell clones were then isolated by a limiting dilution single cell cloning in 96-well plates
提取方案:
DissociateCells were washed, trypsinized, passed through a 40μM cell strainer, centrifuged at 400g, and resuspended at a concentration of 1,000 cells/μL in PBS + 0.5% BSA.Single cells were processed through the Chromium Single Cell 3′ Solution platform using the Chromium Single Cell 3’ Gel Bead, Chip and Library Kits (10X Genomics), as per the manufacturer’s protocol.
建库方案:
-
测序信息
分子类型:
poly(A)+ RNA
库的片段类型:
PAIRED
库的链类型:
Forward
测序平台:
ILLUMINA
测序仪型号:
Illumina HiSeq 2500
链特异性:
Specific
样本
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数据来源
GEN样本编号
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组织
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Condition Detail
生长方案
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分子类型
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加标（Spike-In）
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文章
Genetic and transcriptional evolution alters cancer cell line drug response.
