μCaler RP Solution (for Illumina®) NadPrep RNA & DNA Library Co-Preparation Module NadPrep Universal Stubby Adapter (UDI) Module μCaler Hybrid Capture Reagents v2 μCaler NanoBlockers (for Illumina®) μCaler RP Panel v1.0 Thoroughly Read This Manual Before Operation. For Research Use Only. Version

For research use only. Not for use in diagnostic procedures. This instruction is intended for use with the μCaler RP Solution (for Illumina®). Nanodigmbio Pte. Ltd. reserves all rights.

• Physically separate the laboratory space and establish an operating area for RNA handling, with disinfecting regularly. All the equipment and consumables are dedicated. • Gently pipet RNA rather than thoroughly vortex. • Place the RNA specimens on ice and operate immediately once outside the freezer to avoid degradation. • Make sure to proceed the experiment from RNA handling to 2nd strand synthesis by using RNase-free consumables and reagents. Gloves are essential around all steps. Replace new gloves once contacting the equipment or space out of the RNase-free area. • The personal protective...

μCaler Respiratory Pathogen Solution (referred to as μCaler RP Solution) integrates the RNA & DNA Library CoPreparation Kit, the exclusive patented μCaler Hybrid System, and the XCapViz Bioinformatics Analysis Visual System. It enables a seamless "sample in, result out" workflow within a single day. This solution is designed to cover over 99% of common respiratory pathogens, including viruses, bacteria, fungi, mycoplasma, and chlamydia. It provides comprehensive support for multiplex respiratory pathogen co-detection, precise identification and typing, as well as variants tracking of influenza...

^Caler RP Solution (for Illumina®) consists of the RNA & DNA Library Co-Preparation Module, Adapter Module, |iCaler NanoBlockers (for Illumina®), |iCaler Hybrid Capture Reagents, and |iCaler RP Panel. The composition of the modules and their components are as follows: NadPrep RNA & DNA Library Co-Preparation Module Catalog# Item Package/Storage 1002411 NadPrep RNA & DNA Library Co-Preparation Module, 24 rxn NadPrep RNA & DNA Library Co-Preparation Module, 96 rxn Package# Component Volume Volume Storage Box1 Co-Random Primer 1st Co-Strand Synth. Buffer 1st Co-Strand Synth. Enzyme 2nd Co-Strand...

NadPrep Universal Stubby Adapter (UDI) Module Catalog# Item NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter (UDI) Module Set B2, 96 rxn NadPrep Universal Stubby Adapter (UDI) Module Set E1, 96 rxn NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby Adapter NadPrep Universal Stubby...

1106102 |iCaler NanoBlockers (for Illumina®), 16 rxn 40 |iL 4 ~ 15°C -25 ~-15°C 1106101 |iCaler NanoBlockers (for Illumina®), 96 rxn 210 |iL 4 ~ 15°C -25 ~-15°C 1105202 |iCaler Hybrid Capture Reagents v2, 16 rxn 1105201 iCaler Hybrid Capture Reagents v2, 96 rxn Package# Component Wash Buffer A Pro Wash Buffer B ^Caler RP PanelCatalog# Item Volume Package/Storage 1101431 pCaler RP Panel v1.0, 96 rxn 210 pL -25 ~ -15°C

Digital electrophoresis Agilent 2100 Electrophoresis Bioanalyzer® system (Cat # G2939AA) Agilent 2200 TapeStation Nucleic Acid system (Cat # G2965AA) Bioptic Qsep100 capillary gel electrophoresis system or equivalent Pipettor General laboratory supplier Thermal cycler General laboratory supplier Benchtop centrifuge General laboratory supplier Microcentrifuge General laboratory supplier Vortex mixer General laboratory supplier Magnet stand Tube protocol: Thermo Fisher DynaMag™ -2 Magnet (Cat # 12321D) Thermo Fisher DynaMag™ -96 Side Magnet (Cat # 12331D) or equivalent Qubit fluorometer Thermo...

Workflow RNA & DNA Duration RNA & DNA mixture or RNA RNA Degeneration & Random Primer Fragmentation/End Repair & A-tailing (RNA & DNA mixture or DNA) Adapter Ligation Post-ligation Cleanup Pre-library Amplification Post-amplification Cleanup Perform Hybridization Perform Capture and Elution Purify and Quantify Library ^Caler Hybrid Capture NadPrep Universal Stubby Adapter Safe Stopping Point

Input Amount: 10-100 ng RNA and 1-100 ng DNA mixed sample. It is recommended to quantify nucleic acid using the Qubit™ 3.0 Fluorometer. If RNA is less than 10 ng, add Carrier RNA to a total input amount of 10 ng. Step 1: RNA Degeneration & Random Primer 1. Thaw Co-Random Primer on ice. Thoroughly vortex and briefly centrifuge to collect contents. 2. Set up each reaction in a 0.2 mL PCR tube on ice as follows: O Note: Choose the appropriate nucleic acid sample (RNA & DNA mixture or RNA) according to the type of extractions. If the RNA & DNA mixture or RNA input is less than 12 pL, add Nuclease...

1. Thaw 2nd Co-Strand Synth. Buffer and 2nd Co-Strand Synth. Enzyme on ice. Thoroughly vortex and briefly centrifuge to collect contents. 2. Set up each reaction in a 0.2 mL PCR tube on ice as follows: 2nd Co-Strand Synth. Buffer 8 |iL 2nd Co-Strand Synth. Enzyme 2 |iL 3. Mix thoroughly and then briefly centrifuge to collect contents. 4. Incubate the tube in the thermal cycler programmed as follows. Make sure the thermal cycler is stabilized at 16°C before loading: o Note: The heated lid should be left open. 1. Thaw Co-FERA Buffer at room temperature. Mix thoroughly and centrifuge to collect...

Step 5: Adapter Ligation 1. Thaw Ligation Buffer at room temperature, mix thoroughly and centrifuge to collect the contents. Keep the tube on ice. o Note: The Ligation Buffer is viscous. Please pipette slowly and carefully to ensure the expected volume. 2. Thaw DNA Ligase on ice. Briefly vortex and centrifuge to collect the contents. 3. Remove the tube in Step 4 from the thermal cycler and set up each reaction on ice as follows: NadPrep Universal Stubby Adapter 2 |iL O Note1: Make sure to add NadPrep Universal Stubby Adapter before other reagents to avoid the adapter self-ligation. O Note2: If...