Additional Reagents and Apparatus for Sample Preparation:
- For General RNA Preparation *
Name Recommended Manufacturer Catalog # Purpose AllPrep RNA/DNA Micro Kit QIAGEN 80284 RNA isolation from cells or tissues RecoverAll™ Total Nucleic Acid Isolation Kit for FFPE Thermo Fisher AM1995 RNA isolation from FFPE tissues RNA 6000 Pico Kit Agilent 5067-1511 RNA quality control calculation High Sensitivity RNA Analysis Kit Agilent DNF-472-1000 RNA quality control calculation
- For DirectCell DriverMap Analysis *
Name Recommended Manufacturer Catalog # Purpose TurboCapture 96 or 384 mRNA Kit QIAGEN 72251/72271 Direct use with cell lysate Buffer TCL, 2× (8.5 ml) QIAGEN 1031586 Cell lysis
Additional Reagents and Apparatus for cDNA Synthesis and Amplification:
| Name | Recommended Manufacturer | Catalog # | Purpose |
|---|---|---|---|
| Agencourt® AMPure® XP (Magnetic Beads) | Beckman-Coulter | A63881 | PCR product purification |
| Dynabeads® MPC®-S (Magnetic Particle Concentrator) for 1.5 ml Test Tubes | Thermo Fisher | A13346 | PCR product purification |
| Semi-skirted 96-well PCR plates | Eppendorf | 951020303 | RT-PCR amplification |
| Clear Adhesive Film for 96-well plates | Bio-Rad | MSB1001 | PCR plate seal |
| High Sensitivity DNA Kit | Agilent | 5067-4626 | PCR product QC/quantification |
| High Sensitivity NGS Fragment Analysis Kit (1 bp – 6000 bp) | Agilent | DNF-474-1000 | PCR product QC/quantification |
| Qubit® dsDNA HS (High Sensitivity) Assay Kit | Thermo Fisher | Q32851 | Quantify Indexed Libraries for NGS |
| NextSeq® 500/550 High Output Kit v2 (75 cycles) or HiSeq®/MiSeq® equivalent | Illumina | FC-404-2005 (NextSeq) | Next-Gen Sequencing |
Required Instrumentation:
The protocols were optimized using the specific instruments specified below. Some modifications and optimization to the protocol may be necessary if using different models or instruments from other manufacturers.
| Description | Model | Manufacturer |
|---|---|---|
| Spectrophotometer (NanoDrop or equivalent) |
NanoDrop® ND-2000 | Thermo Fisher |
| Thermal Cycler (Veriti 96-Well or equivalent) |
Veriti® 96-Well | Applied Biosystems |
| Instrument or Gel System to Analyze DNA Fragmentation |
2100 Bioanalyzer® | Agilent |
| Fragment Analyzer™ | Agilent | |
| Standard gel imaging system | many suppliers | |
| DNA Quantification | Qubit 3.0 Fluorometer | Thermo Fisher |
| Next-Generation Sequencer | NextSeq, HiSeq, or MiSeq | Illumina |
Other than the specific reagents and instruments listed above, the protocols assume the user has access to standard materials (e.g., polypropylene tubes, pipette tips), equipment (table top centrifuges, pipettes, scales), and common reagents (e.g., TE buffer, ethanol) and buffers used in a typical life science laboratory.
Last modified:
4 October 2022
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