pLenti-UBC-sgRNA

Cat. No.: P751476

Specifications

Product Name

pLenti-UBC-sgRNA

Product Line

Prodotti biologici

Stoccaggio e spedizione

Condizioni di conservazione di stoccaggio

Store at -80°C

Spedito in

Dry ice packs + Cold packs

Stabilità e conservazione

Stored at -20 ℃, valid for at least 6 months;Stored at -80 ℃, valid for at least one year.

Application Protocols

Tested applications, methods, or recommended conditions are not specified for this item; refer to the CoA/Spec Sheet and your institutional SOPs.

General note: Because protocols depend on the exact backbone features (selection marker, cloning sites, promoters) and the chosen cell system, users should consult the vector map and adopt validated laboratory procedures appropriate to their facility and biosafety level.

Biological Roles

Item-specific biological targets are not specified for this product.

General context (literature):

• sgRNA (single guide RNA) directs CRISPR-associated nucleases (e.g., Cas9) to complementary genomic sequences through base pairing, enabling targeted genome editing, regulation, or imaging when paired with catalytically inactive variants (dCas).
• UBC promoter: the human ubiquitin C promoter is widely used for constitutive expression across many mammalian cell types. In some vector designs, UBC may drive a selection marker or a protein payload; sgRNA expression is commonly under a Pol III promoter (e.g., U6 or H1). The exact arrangement in this item is not specified.
• Lentiviral backbones allow genomic integration of delivered cassettes in transduced cells, enabling stable expression; integration behavior and titer depend on packaging and cell type (not specified for this item).

Biochemical considerations:

• Off-target potential is influenced by spacer sequence, chromatin state, and nuclease variant selection.
• Expression levels driven by the promoter and vector copy number/or integration site affect editing efficiency.

Buffer Applications

This product is a plasmid DNA reagent rather than a conventional buffering agent.

General handling buffers (literature):

• Short-term work: nuclease‑free water or low-salt Tris buffer; maintain pH near neutral (pH 7.5–8.5) to preserve DNA integrity.
• Long-term storage: TE buffer (10 mM Tris-Cl, 1 mM EDTA, pH 8.0) is commonly used to chelate divalent cations and suppress nuclease activity.
• Enzymatic reactions: consult enzyme manufacturer; many assemblies and digestions prefer low-EDTA or EDTA-free conditions.

Electrophoresis and analysis (literature):

• Use standard TAE/TBE buffers for agarose gel analysis of plasmid topology and insert verification. Not an application of the product as a buffer, but relevant to its analysis.

Green Alternatives

As a plasmid DNA reagent, green considerations focus on workflow choices rather than solvent substitution.

Greener choices and trade-offs (literature):

• Direct RNP delivery: Use Cas9/Cas12 ribonucleoprotein with chemically synthesized sgRNA instead of plasmid or lentiviral delivery.
  • Pros: transient activity, reduced off-target duration, avoids viral packaging.
  • Cons: higher upfront reagent cost; delivery can be less efficient in some cell types.
• Non-viral delivery of plasmids: electroporation or lipid/polymer transfection instead of generating lentiviral particles.
  • Pros: reduces BSL‑2 waste and viral reagents.
  • Cons: lower efficiency in hard-to-transfect or primary cells.
• Minimize plastic and hazardous waste: employ reusable glass where safe, choose DNA purification kits with reduced solvent use, and consolidate prep steps to reduce consumables.
• Enzyme choices: use high-efficiency assembly enzymes to lower reaction volumes and repeats.

Note: If lentiviral packaging is required, scale production appropriately, segregate biohazard streams efficiently, and inactivate waste with approved methods (e.g., bleach) to minimize environmental burden.

Pharmaceutical Uses

Not applicable. This product is designated for research use only and is not approved for diagnostic, therapeutic, or manufacturing use.

General industry context (literature):

• Plasmid DNA may be used in research-stage process development and analytical method development. Transition to clinical or GMP applications requires dedicated GMP-grade materials, validated manufacturing, and regulatory oversight—none of which are specified for this item.

Physical Properties

Item-specific physicochemical specifications are not provided for this product; refer to CoA/Spec Sheet.

General/literature properties for plasmid DNA materials:

• Physical state: supplied as aqueous solution or lyophilized DNA depending on configuration; exact appearance for this item not specified.
• Solubility: plasmid DNA is water‑soluble; typically stored in nuclease‑free water or TE (10 mM Tris-Cl, 1 mM EDTA, pH 7.5–8.5) (literature).
• pH of storage buffers: commonly pH 7.5–8.5 (literature). Not specified for this item.
• UV absorbance: A260 for concentration determination; A260/A280 ~1.8–2.0 indicates low protein contamination (literature). Not a specification for this item.
• Thermal properties: DNA denatures above ~90–95°C, but plasmid integrity is best preserved by avoiding repeated freeze–thaw (literature).
• Density, melting point, boiling point, refractive index: not applicable/meaningful for macromolecular DNA preparations.

Practical notes (literature):

• Avoid divalent cation contamination (Mg2+, Ca2+) if nuclease activity is a concern.
• For long-term integrity, store at ≤ −20°C (short term) or −80°C (long term) in low‑EDTA buffers; item-specific storage for this product is −80°C (see Storage & Reconstitution).

Quality and Grades

Item-specific grade, purity, and QC metrics are not specified for this item; refer to CoA/Spec Sheet.

General context for plasmid DNA quality (literature/industry practice):

• Sequence identity: vectors used for cloning or expression are typically sequence-verified across key features (promoters, sgRNA scaffold, selection markers). Verification status for this item is not specified.
• Endotoxin level: for cell culture or transfection, low-endotoxin preparations are often preferred; specific endotoxin specification for this item is not provided.
• Topology: supercoiled fraction influences transfection efficiency. Not specified for this product.
• Residual contaminants: proteins, salts, genomic DNA, and RNase/DNase are common QC targets; acceptance criteria are not specified here.

Grades explained (general):

• Research-use only (RUO): intended solely for laboratory research, not for diagnostic or therapeutic applications.
• “Transfection-grade” or “endotoxin-free” (when applicable) indicate purification optimized for mammalian cell transfection; such designations are not stated for this listing.

Recommendation: Consult the item’s CoA/sequence map for antibiotic resistance marker, backbone size, promoter orientation, and any selectable reporters to ensure compatibility with your host strain and workflow.

Reaction and Applications

Manufacturer applications are not specified; the following are general use-cases for plasmid vectors designated “pLenti‑…‑sgRNA” (literature; verify against this item’s actual map):

• Cloning of sgRNA spacers: insertion of 20 nt target spacers into a sgRNA scaffold via Golden Gate (Type IIS) or Gibson assembly. Orientation and overhangs depend on the specific vector design (not specified here).
• Mammalian expression of sgRNA: delivery of the sgRNA cassette into cells by transfection, electroporation, or via lentiviral packaging and transduction when used with compatible packaging plasmids (institutional biosafety approvals required).
• Multi-component CRISPR workflows: pairing with a Cas9/Cas variant expressed from a separate vector or a stable cell line. UBC promoter (if present) typically drives broad expression of a protein or marker; exact role in this construct is not specified.
• Library construction: adaptation for pooled sgRNA libraries using barcoded oligos (vector specifics required).

Practical tips (general):

• Verify sequence and spacer insertion by Sanger or NGS prior to functional assays.
• Maintain supercoiled topology for maximal transfection efficiency.
• For lentiviral production, use 3rd/4th gen packaging systems and include appropriate controls; perform RCL testing as mandated.

Reaction Conditions

No item-specific reaction conditions are provided.

General conditions (literature) relevant to sgRNA vector workflows:

• Golden Gate cloning: 37°C digestion/ligation cycling with Type IIS enzyme (e.g., 37°C/16°C cycles for 5–30 cycles), small reaction volumes (10–25 µL), and molar insert:vector ratios typically 3–10:1. Exact enzymes and cycling programs should follow enzyme manufacturer guidelines.
• Gibson assembly: isothermal 50°C for 15–60 min; overlaps 20–40 bp; high-fidelity polymerase recommended for fragment generation.
• Transformation: chemically competent E. coli at 42°C heat shock or electroporation per competent-cell specifications; recover in SOC and plate on appropriate antibiotic selection (antibiotic not specified for this item).
• Transfection (non-viral): reagent- and cell-type–dependent; optimize DNA amount and cell density; serum compatibility varies by reagent.
• Lentiviral production: requires compatible packaging plasmids and BSL‑2 practices; follow institutional SOPs. Avoid providing stepwise protocols; consult institutional biosafety documentation.

Always verify the vector map to adjust conditions (restriction sites, antibiotic selection, insert orientation) to the specific construct.

Safety and Handling

Hazard statements, pictograms, and GHS classification are not specified for this item; consult the SDS for authoritative safety information.

General guidance for recombinant plasmid DNA (literature/best practice):

• Biosafety: handle under institutional biosafety guidelines for recombinant DNA (typically BSL‑1 for non-pathogenic E. coli plasmids; institutional assessment required). If used for lentiviral particle production, follow BSL‑2/2+ procedures as mandated by your IBC.
• PPE: lab coat, gloves (nitrile), and eye protection when handling solutions and during transfection/packaging workflows.
• Incompatibilities: avoid nucleases (RNase/DNase) and microbial contamination; maintain aseptic, nuclease‑free technique.
• First aid (general):
  • Skin/eye contact with buffers: rinse with water. Seek medical attention if irritation persists.
  • Ingestion/inhalation: unlikely exposure in normal lab use; follow institutional response procedures.
• Special risks: lentiviral production introduces additional risks (replication-competent lentivirus screening, sharps control, biological waste deactivation via 10% bleach or approved disinfectants).

Waste disposal:

• Dispose of plasmid-containing materials as biohazardous waste per local regulations. Decontaminate surfaces with appropriate DNA/RNA decontamination solutions or freshly prepared bleach.

Solvent Selection

Not a classical solvent; this product is a plasmid DNA reagent typically supplied in an aqueous buffer.

General guidance (literature):

• Preferred diluents: nuclease‑free water or TE buffer (10 mM Tris-Cl, 1 mM EDTA, pH 7.5–8.5) to stabilize DNA and chelate trace metals that can support nuclease activity.
• Avoid: repeated freeze–thaw in pure water; prolonged storage at acidic pH; buffers containing divalent cations if nuclease contamination is a risk.
• Compatibility: suitable for downstream enzymatic steps (restriction digestion, Gibson/Golden Gate assembly) when diluted in low‑salt, nuclease‑free solutions.

Comparison (literature):

• Water (nuclease‑free): optimal for short-term work and enzymatic reactions where EDTA could inhibit enzymes.
• TE buffer: superior for long-term storage stability; EDTA may inhibit downstream enzymes unless diluted.

Storage and Reconstitution

Item-specific facts from Product Data:

• Storage Conditions: Store at −80°C.
• Shipped In: Dry ice packs + Cold packs.

General/literature guidance for plasmid DNA handling (not specifications):

• Upon receipt: keep frozen; minimize time at ambient temperature. If lyophilized, briefly centrifuge the vial before opening to collect contents.
• Reconstitution (if provided dry): use nuclease‑free water or TE buffer (pH 7.5–8.5). Typical concentration targets are user‑defined; mix gently by pipetting—avoid vigorous vortexing.
• Aliquoting: prepare single‑use aliquots to avoid repeated freeze–thaw cycles which can nick plasmid DNA and reduce transfection performance.
• Short-term storage: ≤ −20°C; Long-term storage: −80°C as specified for this item. Avoid frost‑free freezers that cycle temperatures.
• Thawing: thaw on ice, keep solutions cold, and immediately return unused material to −80°C.
• Nuclease control: use sterile, nuclease‑free consumables. Store in low-adsorption tubes to minimize DNA loss at low concentrations.

For exact formulation (buffer composition, concentration) and any stabilizers, consult the CoA/Spec Sheet for this SKU.

Structure and Identity

Item-specific identifiers (CAS, InChIKey, SMILES, exact length) are not specified for this item; refer to CoA/Spec Sheet.

General/literature description:

• pLenti-UBC-sgRNA denotes a recombinant plasmid based on a lentiviral backbone designed for expression of a guide RNA (sgRNA) cassette. “UBC” commonly refers to the human ubiquitin C promoter, a broadly active mammalian promoter (literature). Specific promoter and cassette architecture for this item are not specified.
• Molecular entity: circular, double‑stranded DNA plasmid. No defined small‑molecule formula/structure applies.
• Typical structural features in such vectors (literature; may vary):
  • Lentiviral LTR elements and packaging signals (e.g., 5'LTR, Ψ, RRE, cPPT, 3'LTR/ΔU3) for third‑generation packaging systems.
  • A U6 or other Pol III promoter upstream of a sgRNA scaffold; “UBC” promoter may drive an additional marker or component depending on design.
  • Bacterial origin of replication (e.g., ColE1) and an antibiotic resistance marker for E. coli propagation.
  • Multiple cloning site (MCS) or Golden Gate-type cloning site for sgRNA spacer insertion.
• Functional groups: nucleic acid polymer with phosphodiester backbone; no stereocenters beyond native nucleic acid chirality.

Note: Exact sequence map, length (bp), antibiotic marker, and cloning sites are not provided here; consult the vector map/sequence file on the CoA/Spec Sheet.

Synthetic Utility

In the context of molecular biology, the “synthetic utility” pertains to the vector’s role in assembling genetic constructs.

General/literature utility for pLenti‑style sgRNA vectors:

• Modular cloning: insertion of sgRNA spacer via Golden Gate (Type IIS, e.g., BsmBI/BbsI) or Gibson/SLiCE assemblies. The specific restriction sites for this item are not specified.
• Payload integration: ability to combine sgRNA expression with additional cassettes (e.g., selectable markers, reporters) through standard cloning workflows.
• Retrosynthetic planning (genetic): decompose desired function into promoter, spacer, scaffold, terminator, and selection modules; ensure non-conflicting restriction sites and reading frames where applicable.
• Library assembly: supports pooled oligo cloning for gRNA libraries when backbone architecture allows.

Note: Verify backbone antibiotic resistance, origin of replication, and any lentiviral elements in the provided sequence map for compatibility with host strains and packaging systems.

Target Specificity

Not specified for this item; refer to CoA/Spec Sheet.

Note: For sgRNA vectors, “target specificity” is determined by the 20 nt spacer sequence inserted by the user. This listing does not provide a defined spacer or target gene information.

Domande frequenti

How should this product be stored?

Store at ?80 °C. Ultra-low temperature storage is required to maintain the specified shelf life.

How is this product shipped?

This product ships frozen with dry ice and cold packs. Unpack on arrival and transfer it to the storage condition stated above; handle dry ice with insulated gloves in a ventilated area.

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.

Which species reactivity has this antibody been validated against?

Validated reactivity is listed on the PDP and on each lot's datasheet. We validate against the most commonly tested species for the target (pLenti-UBC-sgRNA); cross-reactivity in untested species may exist but is not guaranteed. Contact technical support for specific species inquiries.