RPAP1 Human Pre-designed siRNA Set A

Cat. No.: R1460124

Storage

Store at -20°C

Shipped In

Ice chest + Ice pads

Panoramica

RPAP1 Human Pre-designed siRNA Set A contains three designed siRNAs for RPAP1 gene (Human), as well as a negative control, a positive control, and a FAM-labeled negative control. Components RPAP1 siRNA-1: 5 nmol (HPLC) RPAP1 siRNA-2: 5 nmol (HPLC) RPAP1 siRNA-3: 5 nmol (HPLC) siRNA Negative Control: 5 nmol (HPLC) FAM-labeled siRNA Negative Control: 5 nmol (HPLC) GAPDH siRNA Positive Control:5 nmol (HPLC)

Specifications

Condizioni di conservazione di stoccaggio

Store at -20°C

Spedito in

Ice chest + Ice pads

Questo prodotto richiede spedizione a catena fredda. I servizi di terra e altri servizi economici non sono disponibili.

Product Line

Prodotti biologici

Application Protocols

General transfection protocol (adherent cells; literature guidance, not item-specific)

1. Preparation: Work RNase-free. Thaw siRNA on ice, briefly spin, and keep cold. Prepare growth medium and transfection reagent per manufacturer guidance.
2. Seeding: Plate cells 18–24 h before transfection to reach ~60% confluence at time of transfection.
3. Complex formation: Dilute siRNA to desired working concentration (e.g., 10–50 nM final) in serum-free medium. In a separate tube, dilute transfection reagent. Combine, gently mix, and incubate 5–20 min at room temperature.
4. Transfection: Add complexes dropwise to cells. Optionally replace with complete medium after 4–6 h if required by reagent; many modern reagents allow no-change protocols.
5. Incubation: Culture 24–96 h. Assess mRNA at 24–48 h and protein at 48–96 h.
6. Analysis: Include non-targeting control, positive control siRNA, and cytotoxicity assessment. Normalize qPCR to multiple housekeeping genes; confirm knockdown by Western blot.

Electroporation (hard-to-transfect cells)

• Resuspend cells in electroporation buffer with siRNA (typical 50–200 nM). Apply program optimized for cell type. Immediately recover in pre-warmed medium.

Notes

• Optimize siRNA dose and reagent ratio per cell line to balance efficacy and viability.
• For this specific product, any validated protocols, sequences, or recommended doses are Not specified; refer to CoA/Spec Sheet.

Biological Roles

Target overview (literature)

• RPAP1 (RNA polymerase II-associated protein 1) is implicated in the assembly and functional regulation of the RNA polymerase II (Pol II) transcriptional machinery. Studies indicate roles in facilitating the interaction of Pol II with general transcription factors and/or Mediator, contributing to initiation competence and promoter engagement.
• RPAP1 has been connected to proper Pol II complex integrity and may influence CTD (C-terminal domain) phosphorylation states through recruitment dynamics of regulatory factors (literature context).

Relevance of siRNA-mediated knockdown (general)

• Depleting RPAP1 allows dissection of transcriptional control at early stages, mapping dependency of promoter-proximal pausing, elongation entry, and coactivator engagement.
• Transcriptomic profiling (RNA-seq) post-knockdown can identify RPAP1-dependent gene sets, while ChIP-qPCR/ChIP-seq can examine Pol II occupancy shifts.
• Functional assays might include measurements of nascent RNA synthesis (e.g., EU labeling), reporter assays for promoter activity, and analysis of global RNA processing changes.

Caveats and best practices

• Off-targeting and innate immune activation can confound interpretations; incorporate multiple independent siRNAs (as in a set), appropriate non-targeting controls, and, where possible, rescue with an siRNA-insensitive RPAP1 construct.
• Time-resolved sampling (e.g., 24, 48, 72 hours) can distinguish primary transcriptional effects from downstream secondary responses due to altered transcription homeostasis.

Note: The above summarizes literature context; it is not item-specific.

Buffer Applications

Applicability: While this product is not itself a buffer, successful use depends on appropriate RNase-free buffers for reconstitution, storage, and delivery.

Common buffer systems (general guidance)

• RNase-free water: Preferred for initial dissolution and short-term working aliquots. pH near neutral.
• TE/TE-low: 10 mM Tris·Cl, pH 7.5–8.0, 0.1–1 mM EDTA. EDTA chelates Mg2+/Mn2+ to slow hydrolysis and nuclease activity. Use lower EDTA (≤0.5 mM) if forming complexes with cationic lipids/polymers.
• siRNA buffer: Commercial or lab-made (e.g., 10 mM Tris·Cl, 20 mM KCl, pH 7.5) supports stability and compatibility with many transfection reagents.
• Complexation diluents: Follow transfection reagent guidance (often serum-free, reduced ionic strength media like Opti-MEM) for forming lipid:siRNA complexes.

Practical notes

• Sterility and RNase control are paramount; prepare buffers with certified RNase-free reagents and tools.
• Avoid repeated freeze–thaw; prepare small aliquots of reconstituted siRNA in buffer of choice.
• Avoid phosphate or high-salt buffers during complex formation unless specifically supported by the transfection reagent, as ionic strength can interfere with nanoparticle assembly.

Item-specific buffer recommendations, concentrations, and any supplied buffer composition are Not specified for this item; refer to CoA/Spec Sheet.

Green Alternatives

Applicability note: This is a nucleic acid reagent used in cell-based assays. Green considerations center on delivery chemistry, plastic use, and waste minimization rather than solvent substitution.

Greener practice options (general)

• Transfection reagents: Prefer lower-toxicity, serum-compatible lipid formulations to reduce media exchanges and aqueous waste. Evaluate biodegradable polymers over PEI when feasible.
• Process minimization: Use miniaturized formats (96/384-well) and automation to lower reagent volumes, plastic consumption, and energy for incubation.
• Buffer preparation: Make concentrated stock buffers to reduce packaging; use reusable glass where RNase control permits. Prepare only what is needed to limit disposals.
• Cold-chain efficiency: Consolidate shipments and coordinate deliveries to reduce dry-ice/ice-pack waste and transport emissions when institutional logistics allow.

Comparison (general, illustrative)

• Lipid-based delivery: Lower energy input, compatible with many lines; generates plastic consumable waste; often fewer wash steps.
• Electroporation: No chemical carriers; higher energy demand per run; may require larger cell numbers and cuvettes but avoids persistent polymers.

Note: The product formulation, if any, is Not specified for this item; refer to CoA/Spec Sheet and your institution’s sustainability guidelines for implementation details.

Pharmaceutical Uses

Scope limitation: This product is labeled For research use only and is not intended for human or veterinary use, diagnostic procedures, or as a pharmaceutical ingredient.

General context (non-clinical, literature)

• Research-grade siRNAs are widely used in preclinical discovery to identify and validate genetic targets, characterize mechanism of action, and inform small-molecule or biologic development strategies.
• In formulation research (lab-scale), siRNAs may be encapsulated in lipid nanoparticles or complexed with cationic polymers to study delivery performance in cell culture models. Such experiments guide selection of delivery chemistries in discovery workflows, not for clinical application.
• Assay development: siRNA knockdown can serve as a benchmark for target engagement assays, helping define dynamic range and expected phenotypic outcomes prior to screening chemical libraries.

Quality/regulatory note

• GMP status, residual solvents, endotoxin, and sterility requirements for clinical materials are substantially more stringent than for research reagents. For this item, no pharmaceutical grade or GMP claims are made; any specific compliance information is Not specified for this item; refer to CoA/Spec Sheet.

Conclusion

• Use this siRNA set solely for laboratory research. Do not incorporate into therapeutic formulations or in vivo studies intended for clinical progression without appropriate grade, documentation, and approvals.

Physical Properties

Item-specific properties

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular weight (per duplex) and exact sequence length: Not specified for this item; refer to CoA/Spec Sheet.
• Extinction coefficient (ε260) and OD-to-mass conversion: Not specified for this item; refer to CoA/Spec Sheet.

General/literature properties for siRNA (not item-specific)

• Physical state: Typically supplied lyophilized or in aqueous, RNase-free solution at low millimolar to micromolar concentrations.
• Solubility: Freely soluble in RNase-free water or low-salt buffers (e.g., 1× siRNA buffer, 10 mM Tris·Cl, pH ~7.4–8.0). Highly polar and hydrophilic; insoluble in organic solvents.
• UV characteristics: Strong absorbance near 260 nm; A260 is used for quantitation. Duplex formation slightly hypochromic relative to single strands.
• pKa/logP: Not generally applicable; macromolecular polyanions with multiple ionizable phosphate groups remain negatively charged at physiological pH.
• Melting temperature (Tm): Dependent on sequence, length, and ionic strength; typical siRNA duplexes exhibit Tm values often in the 60–85 °C range in standard buffers (literature, sequence-dependent).
• Stability: Sensitive to RNases; chemically stable when dry and stored frozen. In solution, stable for weeks at −20 °C and for months at −80 °C; repeated freeze–thaw can reduce integrity (literature guidance).

Note: Exact numeric values for this specific set are Not specified for this item; refer to CoA/Spec Sheet.

Quality and Grades

Item-specific information

• Grade/purity: Not specified for this item; refer to CoA/Spec Sheet.
• Stabilizers/modifications: Not specified for this item; refer to CoA/Spec Sheet.

General notes on siRNA quality (literature/industry practice)

• Purity levels: Common offerings include desalted (crude), cartridge-purified, or HPLC/ PAGE-purified duplexes. Higher-purity materials typically reduce off-target effects from truncated/failed sequences and improve reproducibility in sensitive assays.
• Identity and QC: Typical QC includes HPLC traces, LC–MS of single strands, OD260-based quantification, and annealing confirmation. Stringent QC is valuable for transfection into difficult cell types or for in vivo-like delivery studies (research-only here).
• Endotoxin/bioburden: For cell culture, low endotoxin is preferred; some vendors report EU levels. For this item, any such specifications are Not specified; refer to CoA/Spec Sheet.
• UV transparency/OD reporting: OD260 units are often converted to nmol/mg using sequence-specific extinction coefficients. Without sequences, exact conversions are not applicable here.
• Impact of chemical modifications: 2′-O-Me/2′-F sugars and phosphorothioate linkages can enhance nuclease resistance and duplex stability, potentially lowering required dose. Whether such modifications are present in this product is Not specified; refer to CoA/Spec Sheet.
• Lot-to-lot consistency: For multi-duplex sets, consistency in synthesis scale, purification method, and annealing protocol supports reproducible knockdown benchmarking across duplexes.

Reaction and Applications

Context: As a functional genomics reagent, this siRNA set is designed to downregulate human RPAP1 expression via RNA interference (RNAi). There are no classical organic reactions here; rather, cellular processes are engaged.

Applications (general, not item-specific)

• Gene knockdown: Transient suppression of RPAP1 mRNA to interrogate Pol II-related transcriptional mechanisms, cofactor dependencies, or pathway mapping.
• Phenotypic screening: Use multiple duplexes (as in a Set A) to mitigate off-target risk; consensus phenotypes across duplexes strengthen target attribution.
• Mechanistic studies: Time-course knockdown to examine primary vs secondary transcriptional effects; assess RNA Pol II occupancy or CTD phosphorylation states by ChIP-qPCR or Western blot.
• Synthetic lethality/interaction mapping: Combine RPAP1 knockdown with chemical perturbations (e.g., transcriptional inhibitors) to probe pathway robustness.

Practical considerations

• Transfection modality: Lipid-based (adherent cells), polymeric, or electroporation (primary cells/suspension lines). Optimize dose (e.g., 5–50 nM siRNA) per cell type and reagent (literature ranges; not item-specific).
• Controls: Non-targeting siRNA, positive knockdown control, and siGLO/fluorescent control where available. Rescue experiments with siRNA-resistant RPAP1 cDNA bolster specificity claims.
• Readouts: qRT-PCR for mRNA (24–48 h), immunoblot for protein (48–96 h), functional assays (e.g., nascent transcription, cell cycle).

Item-specific sequence details and recommended working concentrations are Not specified for this item; refer to CoA/Spec Sheet.

Reaction Conditions

Context: Here “reaction conditions” refer to cell-based transfection and knockdown parameters rather than chemical synthesis.

General transfection conditions (literature guidance; not item-specific)

• Cell seeding: Plate cells to reach ~50–70% confluence at transfection. Suspension/primary cells may require electroporation.
• Dose: Typical starting range 5–50 nM total siRNA per well; titrate to minimize off-target effects while achieving knockdown.
• Complexation: Mix siRNA with transfection reagent in serum-free medium; incubate 5–20 min to form complexes; then add to cells. Many modern reagents permit direct addition in serum-containing media—follow vendor instructions.
• Incubation: 24–72 h for mRNA knockdown assessment by qRT-PCR; 48–96 h for protein level reduction by Western blot/IF, depending on protein half-life.
• Controls: Non-targeting siRNA, positive control siRNA, and (optionally) siRNA-resistant RPAP1 expression plasmid for rescue.
• Temperatures: Standard mammalian culture (37 °C, 5% CO2). Avoid temperature shocks during complex formation.
• Replicates: Perform biological triplicates with technical duplicates for robust statistics.

Expected outcomes (typical literature ranges)

• Effective duplexes can achieve >70% mRNA knockdown under optimized conditions; protein suppression depends on turnover. Actual performance varies by cell type and delivery reagent.

Note: Specific recommended conditions for this product are Not specified; consult CoA/Spec Sheet and your transfection reagent documentation.

Safety and Handling

Item-specific safety information

• GHS classification, signal word, H-statements, pictograms: Not specified for this item; refer to SDS.
• Research use note: For research use only.
• Storage condition (from Product Data): Store at −20 °C. Shipped in ice chest + ice pads.

General safety guidance for siRNA reagents (literature/best practice)

• Hazard profile: Synthetic siRNAs are generally considered low hazard chemical/biological reagents but should be handled using standard laboratory PPE (lab coat, gloves, eye protection) and good microbiological practices.
• RNase control: Work in a clean, RNase-free environment. Use RNase-free tubes, barrier tips, and freshly opened gloves. Avoid touching tube rims; wipe benches and tools with RNase decontaminants.
• Incompatibilities: Avoid RNase contamination and prolonged exposure to elevated temperatures. Avoid repeated freeze–thaw cycles; aliquot upon first thaw.
• First aid (general): In case of skin or eye contact, rinse with water. If inhaled (aerosols/powders), move to fresh air. If ingested, rinse mouth. Seek medical advice if symptoms persist. Refer to SDS for authoritative instructions.
• Spill response: For solutions, wipe with RNase inactivation solution followed by water. For dried material, gently dampen to avoid aerosolization, then clean per institutional procedures.
• Waste: Dispose of in accordance with local regulations for non-hazardous laboratory biotech waste unless institutional policies specify otherwise.

Always consult the product-specific SDS and institutional EHS guidance before use.

Solvent Selection

Applicability note: This product is a nucleic acid reagent rather than an organic solvent. Solvent selection pertains to reconstitution and handling buffers.

Recommended media (general, not item-specific)

• Primary: RNase-free water (molecular biology grade) for initial dissolution; suitable for short-term storage and most transfections.
• Stabilizing buffer: 1× siRNA buffer or TE/TEpH (e.g., 10 mM Tris·Cl, pH 7.5–8.0, 0.1–1 mM EDTA) to chelate trace divalent cations that catalyze RNA degradation. Avoid high EDTA if using cation-dependent transfection reagents.
• Physiological buffers: For direct complexation prior to delivery, many lipid-based transfection agents prefer serum-free, low-ionic-strength buffers (e.g., Opti-MEM or vendor-specified diluent) during complex formation.

Miscibility and polarity (literature)

• siRNA is highly polar and polyanionic; insoluble in organic solvents. Avoid alcohols/organic co-solvents during complexation as they can disrupt duplex integrity or transfection reagent performance.

Practical tips

• Bring solutions to room temperature before pipetting to improve accuracy.
• Prepare working stocks (e.g., 10–20 µM) in RNase-free water or buffer; verify compatibility with the chosen transfection reagent.
• Filter-sterilization is generally unnecessary if prepared with sterile materials; avoid 0.2 µm filtration of dilute siRNA to prevent adsorption losses.

Item-specific solvent guidance is Not specified for this item; refer to CoA/Spec Sheet and your transfection reagent manufacturer’s recommendations.

Storage and Reconstitution

Item-specific storage

• Storage condition: Store at −20 °C (from Product Data).
• Shipping: Shipped in ice chest + ice pads (from Product Data).

General reconstitution guidance (not item-specific)

• Upon receipt: If lyophilized, briefly centrifuge to collect material. Reconstitute in RNase-free water or buffer (e.g., 10 mM Tris·Cl, pH 7.5–8.0, 0.1–0.5 mM EDTA) to a convenient stock (e.g., 20–100 µM). If supplied in solution, verify concentration and buffer on the label/CoA.
• Mixing: Gently pipette up/down; avoid vigorous vortexing that may shear or foam complexes.
• Aliquoting: Prepare single-use aliquots to avoid repeated freeze–thaw cycles. Label with concentration, buffer, and date.
• Storage: Short term (days to weeks) at −20 °C; long term at −80 °C. Protect from repeated freeze–thaw and from RNase contamination. Avoid frost-free freezers that cycle temperatures.
• Thawing: Thaw on ice, briefly spin to collect condensate. Keep on ice during setup; return unused aliquots promptly to −20/−80 °C.
• Stability: In general, dry siRNA is stable for months at −20 °C; in solution, stability improves at −80 °C. Exact stability for this item is Not specified; refer to CoA/Spec Sheet.

Documentation

• Any specific reconstitution volume, final concentration targets, or provided buffer composition for this product are Not specified; refer to CoA/Spec Sheet.

Structure and Identity

Item-specific facts (from Product Data)

• Product: RPAP1 Human Pre-designed siRNA Set A (SKU: R1460124)
• Target gene: Human RPAP1 (RNA polymerase II-associated protein 1)
• CAS: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular formula: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular weight: Not specified for this item; refer to CoA/Spec Sheet.
• SMILES / InChIKey: Not applicable to a mixed siRNA duplex set; Not specified for this item; refer to CoA/Spec Sheet.

General/literature description of structure (not item-specific)

• Typical siRNA reagents are short double-stranded RNAs, ~19–21 base pairs with 2-nt 3′ overhangs (often dTdT), composed of ribonucleotides with a phosphodiester (or modified) backbone.
• Each duplex comprises an antisense (guide) strand designed to be complementary to the target mRNA sequence and a sense (passenger) strand.
• Optional chemical modifications may be included in some products (e.g., 2′-O-methyl, 2′-F, phosphorothioate linkages) to enhance stability and reduce immunostimulation; presence/absence and extent are Not specified for this item; refer to CoA/Spec Sheet.

2D structural features (literature)

• Linear polyanionic biopolymers with repeating ribose-phosphate backbone and heterocyclic nucleobases (A, U, G, C).
• Duplex adopts A-form helical geometry in solution with base pairing (A–U, G–C) and stacking interactions.

Identity confirmation (general practices)

• Sequence identity and purity are typically established by LC-MS or HPLC chromatograms and OD260 quantification. For this item, any specific analytical certificates are Not specified; refer to CoA/Spec Sheet.

Synthetic Utility

Applicability note: This product is a biological reagent (siRNA) and is not typically used as a building block or reagent in classical organic synthesis.

Relevant laboratory utility (general)

• Functional genomics tool: Employed to silence RPAP1 gene expression, enabling mechanistic studies of transcriptional regulation and pathway dependency mapping in cells.
• Assay calibration: Establishes maximum achievable target suppression to calibrate phenotypic assays prior to medicinal chemistry campaigns.
• Genetic interaction studies: Combines with CRISPRi/a, shRNA, or overexpression systems to perform epistasis analysis and network inference.

Non-applicability to chemical synthesis

• No roles in catalysis, protection/deprotection, cross-coupling, or other named reactions.
• No meaningful discussion of solvent polarity parameters, nucleophilicity/electrophilicity, or redox behavior in the context of synthetic transformations.

If your goal is chemical synthesis, consider browsing other catalog sections for relevant reagents (e.g., coupling agents, catalysts, building blocks). For RNA work, this product is best integrated into cell-based assays, transcript analysis workflows, and biochemical characterization of transcriptional complexes.

Target Specificity

Item-specific data

• Intended target: Human RPAP1 (RNA polymerase II-associated protein 1).
• Duplex sequences, seed regions, and any off-target filtering criteria: Not specified for this item; refer to CoA/Spec Sheet.

General considerations for siRNA specificity (literature)

• Primary specificity is governed by perfect complementarity of the antisense (guide) strand to the target mRNA. Seed region (positions 2–8) can mediate microRNA-like off-targeting if partial matches exist in 3′UTRs of unrelated transcripts.
• BLAST or siSPOTR-style computational checks are recommended to evaluate potential off-targets, SNP overlap, and transcript isoform coverage for the chosen cell line/species.
• Use of siRNA pools or sets (multiple independent duplexes) enables cross-validation: Phenotypes shared by multiple duplexes increase confidence in on-target effects.
• Chemical modifications (if present) such as 2′-O-Me in the seed region can reduce off-targeting without greatly compromising efficacy; presence/absence for this product is Not specified; refer to CoA/Spec Sheet.

Validation recommendations

• Quantify RPAP1 mRNA reduction by qRT-PCR with exon–exon junction-spanning primers.
• Confirm protein-level suppression by Western blot using well-validated RPAP1 antibodies and orthogonal methods (e.g., targeted proteomics) when feasible.

Domande frequenti

How should this product be stored?

Store at ?20 °C. Freezer storage is required to maintain the specified shelf life.

How is this product shipped?

This product ships in an insulated container with ice pads. Unpack on arrival and transfer it to the storage condition stated above.

What documentation is provided?

Available product documentation, including Certificates of Analysis (COA), Safety Data Sheets (SDS), and specification sheets, is shown in the product document area. Document availability and access follow the current site policy.