Depending on the type of samples and procedural options followed in the protocol, several of the following items will be required to run the assay.
Neoteryx Mitra Tips with GenTegra RNA-NEO stabilizer for Blood Collection
| No. individuals/ Rxn Kit |
Mitra Tips + GTR-NEO RNA Stabilizer | DriverMap Configuration |
|---|---|---|
| 6 | Qty 1 VM-302SNLR; Mitra 30ul 2 sampler Device 30/PK RUO | 24 rxn |
| 12 | Qty 1 GTRNEO-32; GenTegra GTR-NEO Active Chemical Protection of RNA, 2mL Tube | |
| 24 | Qty 3 VM-302SNLR; Mitra 30ul 2 sampler Device 30/PK RUO | 96 rxn |
| 48 | Qty 3 GTRNEO-32; GenTegra GTR-NEO Active Chemical Protection of RNA, 2mL Tube |
Please inquire for complete sampling collection kits (includes lancets, bandage, gauze, instructions). For more information, visit the Neoteryx Website.
Additional Reagents for Hybridization, cDNA Synthesis, Amplification, and Amplified Libraries QC
| Name | Recommended Manufacturer | Catalog # | Purpose |
|---|---|---|---|
| SPRIselect, 5 ml (Magnetic Beads) | Beckman Coulter | B23317 | RNA-RevGSP hybrid and PCR product purification |
| Dynabeads® MPC®-S (Magnetic Particle Concentrator) for 1.5 ml Test Tubes | Thermo Fisher | A13346 | RNA-RevGSP hybrid product purification in 0.5-1.5-ml test tubes |
| Catalyst 96 slotted ring magnetic plate | Alpaqua | A000550 | RNA-RevGSP hybrid product purification in 96-well plate |
| IGEPAL CA-630 (NP40) | Sigma | 18896 | Detergent additive for hybridization solution in DirectCell protocol |
| 96-well PCR plates | Thermo Fisher | A43673 | RT-PCR reactions |
| Clear Adhesive Film for 96-well plates | Bio-Rad | MSB1001 | PCR plate seal |
| High Sensitivity DNA Kit for Bioanalyzer | Agilent | 5067-4626 | PCR product QC/ quantification |
| High Sensitivity NGS Fragment Analysis Kit (1 bp – 6000 bp) for Fragment Analyzer | 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 Mid Output Kit v2.5 (150 cycles) | Illumina | 20024909 | Next-Gen Sequencing |
| NextSeq® 500/550 High Output Kit v2.5 (150 cycles) | Illumina | 20024908 | Next-Gen Sequencing |
| NextSeq 1000/2000 XLeap-SBS P1 Reagent Kit (100 cycles) | Illumina | 20100983 | Next-Gen Sequencing |
| NextSeq 1000/2000 XLeap-SBS P2 Reagent Kit (100 cycles) | Illumina | 20100987 | Next-Gen Sequencing |
| NextSeq 1000/2000 XLeap-SBS P3 Reagent Kit (100 cycles) | Illumina | 20100990 | Next-Gen Sequencing |
| NextSeq 1000/2000 XLeap-SBS P4 Reagent Kit (100 cycles) | Illumina | 20100994 | Next-Gen Sequencing |
| MiSeq Reagent Kit v3 (150 cycles) | Illumina | MS-102-3001 | Next-Gen Sequencing |
Recommended Instrumentation
This protocol is optimized for the 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 | Thermo Fisher |
| Instrument or Agarose Gel System to analyze DNA fragments or Slab Gel Electrophoresis System to analyze amplified DNA products |
||
| 2100 Bioanalyzer® | Agilent | |
| Fragment Analyzer™ | Agilent | |
| Agarose gel electrophoresis/imaging system | many suppliers | |
| DNA Quantification | Qubit 3.0 Fluorometer | Thermo Fisher |
| Next-Generation Sequencer | NextSeq 2000, NextSeq 500 | Illumina |
| Thermomixer R (for microsampling AIR profiling) | Thermomixer R with heat block for 1.5-ml test tubes | Eppendorf/Fisher Scientific |
Other than the specific reagents and instruments listed above, the protocol assumes the user has access to standard materials (e.g., polypropylene tubes, pipette tips), equipment (tabletop centrifuges, pipettes), and common reagents (e.g., TE buffer, ethanol) and buffers used in a typical life science laboratory.
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