Expression profiling by high throughput sequencing
Summary
The plasticity and heterogeneity of cancer cells are known to play a major role in malignant progression, yet the precise molecular drivers of metastases remain elusive. We performed single-cell RNA-Seq (scRNA-Seq) on tumours and matched lung macrometastases of patient-derived xenografts (PDXs) of breast cancer to identify molecular mechanisms of metastasis during colonization.
Overall design
Tumors and matched lung metastases from PDX models grown in NSG mice were dissociated and human cells purified by FACS . PDX models cells selection relied on the CD298 human-specific marker. Samples derived from a cell line MDA-MB-231 expressing a GFP-Luciferase construct were selected based on GFP expression. Single cell isolation was performed on the Fluidigm C1 platform. mRNA was isolated with the Takara SMART-Seq Ultra Low RNA kit. Libraries for Illumina sequencing were prepared with the Nextera XT DNA Library Preparation Kit.
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