pLenti-CBL-sgRNA

Cat. No.: P744457

Specifications

Product Name

pLenti-CBL-sgRNA

Product Line

Produtos biológicos

Armazenamento e envio

Condições de armazenamento de armazenamento

Store at -80°C

Enviado em

Dry ice packs + Cold packs

Estabilidade e armazenamento

Stored at -20 ℃, valid for 3 months;Stored at -80 ℃, valid for at least one year.

Application Protocols

Only information from Product Data may be listed here.

• Tested/validated applications: Not specified for this item.
• Recommended conditions/dilutions/controls: Not specified for this item.

Note: For general guidance on plasmid transfection or lentiviral packaging, please consult standard literature protocols and your institution’s biosafety procedures. Item-specific protocols, if available, will be provided in the CoA/Spec Sheet.

Biological Roles

General/literature context (non-item-specific)

• sgRNA function in CRISPR: A single guide RNA (sgRNA) provides both target recognition (20-nt spacer) and a structural scaffold that complexes with a CRISPR nuclease (e.g., Cas9). The ribonucleoprotein directs site-specific DNA cleavage, enabling knockout via indels from non-homologous end joining or precise edits via homology-directed repair when a donor is supplied.
• CBL gene background: CBL (Casitas B-lineage lymphoma) encodes an E3 ubiquitin ligase that regulates receptor tyrosine kinase downregulation, endocytosis, and signal attenuation via ubiquitination (literature). Alterations in CBL can modulate pathways such as EGFR and immune receptor signaling and have been implicated in hematologic malignancies (literature). An sgRNA targeting CBL can be used to explore the consequences of CBL loss-of-function on cellular signaling, protein turnover, and proliferative phenotypes in research models.
• Delivery context: In a lentiviral backbone, sgRNAs can be introduced into dividing and non-dividing cells for stable expression (literature). Specific promoters, selection markers, and compatibility with particular Cas9 variants for this item are not specified and should be confirmed from the CoA/Spec Sheet.

Item-specific facts (from Product Data)

• None beyond the product name indicating a CBL-directed sgRNA; no sequence, promoter, or selection details are specified.

Buffer Applications

This product is a plasmid DNA reagent and is not itself a buffer system. Therefore, classical buffer selection (pH ranges, recipes) is not directly applicable.

General/literature practice (non-item-specific)

• Handling buffers: Nuclease-free water or TE buffer (e.g., 10 mM Tris, 1 mM EDTA, pH ~7.5–8.0) are commonly used to store or dilute plasmid DNA. For enzyme reactions (e.g., cloning), use the buffer supplied with the enzyme. For transfection, follow the delivery reagent’s recommended diluent.

Item-specific details

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

Green Alternatives

This is a nucleic-acid research tool rather than a solvent or commodity chemical; classical green-chemistry substitution is not directly applicable.

General/literature considerations to reduce environmental impact (non-item-specific)

• Prefer non-viral delivery when feasible: Electroporation or lipid-mediated plasmid transfection can avoid viral packaging waste and BSL-2 containment, reducing consumables and energy use associated with biosafety cabinets and waste sterilization.
• Right-size prep volumes: Use mini/midi preps commensurate with experimental needs to limit plastic and reagent use.
• Reusable materials: Where permitted, adopt reusable tube racks, ice packs, and minimize single-use plastics. Decontaminate with the least hazardous effective agents.
• Cold-chain efficiency: Consolidate shipments on dry ice to reduce packaging footprint. For this item, shipping is on dry ice packs + cold packs per Product Data.
• Waste management: Segregate biological from chemical waste; neutralize bleach appropriately and follow local regulations.

Pharmaceutical Uses

No pharmaceutical or clinical uses are claimed or supported for this product. It is designated for research use only per Product Data.

General/literature context (non-item-specific)

• Plasmid DNA reagents are widely used in discovery research, target validation, and process development studies. When plasmids are used in manufacturing (e.g., as auxiliary materials for viral vector production), they must meet stringent quality and regulatory criteria (cGMP-grade, comprehensive impurity profiles, and documentation). Such attributes are not specified for this research-use item.

Item-specific facts (from Product Data)

• Research use only. No pharmacopeial status, excipient role, or therapeutic application is specified for this item.

Physical Properties

Item-specific facts (from Product Data)

• Physical state/appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Concentration/formulation: Not specified for this item; refer to CoA/Spec Sheet.

General/literature properties of plasmid DNA (non-item-specific)

• Phase behavior: Macromolecular biopolymer; classical BP/MP are not applicable. Plasmid DNA is handled in aqueous buffers at neutral pH (literature).
• Solubility: Freely soluble in nuclease-free water and common biological buffers (e.g., TE: 10 mM Tris, 1 mM EDTA, pH 7.5–8.0) (literature). Salt, pH, and counterions influence viscosity and stability.
• UV characteristics: DNA exhibits a strong absorbance at 260 nm (A260), with A260/A280 ~1.8 for pure DNA (literature). These are analytical indicators and not item specifications.
• Stability (storage-related): Plasmid DNA is generally stable for long-term storage at −20 to −80°C, protected from nucleases and repeated freeze–thaw cycles (literature). Actual stability for this item is governed by the stated storage condition (−80°C) and the specific formulation (not specified).
• Density, refractive index, logP, pKa: Not applicable/relevant to plasmid DNA solutions; not specified for this item.

Quality and Grades

Item-specific facts (from Product Data)

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

General guidance (non-item-specific)

• Typical QC for plasmid DNA (literature): Sequence verification of the sgRNA spacer and backbone features; restriction digest analysis to confirm plasmid identity; agarose gel electrophoresis to confirm topology and integrity; A260/A280 and A260/A230 ratios for purity; optional endotoxin testing if intended for sensitive cell types.
• Research-use-only status: Labeled “For research use only,” indicating it is not validated for diagnostic, therapeutic, or clinical applications.
• Implications of “pLenti sgRNA” format: Many labs expect compatibility with standard lentiviral packaging systems and sgRNA expression promoters; however, exact backbone, selection marker(s), and any additional cassettes (e.g., reporter, antibiotic resistance) are not specified for this item and must be confirmed by CoA/Spec Sheet.
• Recommendation: Prior to experimental use, verify the plasmid sequence/map, antibiotic selection, and cloning sites against the supplied documentation to ensure compatibility with your cloning strategy, Cas nuclease, and host system.

Reaction and Applications

Item-specific facts (from Product Data)

• Manufacturer applications: Not provided.

General/literature applications for a pLenti sgRNA plasmid (non-item-specific)

• CRISPR gene perturbation: Delivery of an sgRNA targeting the CBL gene to enable Cas9-mediated knockout, nickase-mediated editing, or dCas9-mediated repression/activation when paired with the appropriate Cas nuclease or effector (supplied separately). Spacer sequence and nuclease compatibility for this item are not specified.
• Lentiviral delivery: Use as the transfer vector in third-generation lentiviral systems to package sgRNA for stable integration/transduction of hard-to-transfect cells. Observe BSL-2 practices for viral work.
• Cloning/editing workflows: Swap or confirm the sgRNA spacer via Golden Gate or Gibson Assembly (literature) if redesign is needed. Validate edits by amplicon sequencing, T7E1, or ICE/TIDE analysis.
• Functional genomics: Generate CBL loss-of-function cell models to interrogate ubiquitin ligase pathway biology, signaling, and proteostasis (literature). Downstream readouts may include immunoblotting, phospho-signaling assays, or phenotypic screens.

Practical tips (general)

• Verify plasmid map and sgRNA sequence prior to use. Ensure the sgRNA scaffold and promoter match the Cas nuclease context (SpCas9 vs SaCas9, etc.).
• For transient assays, co-transfect with a Cas9-expression plasmid; for stable lines, consider lentiviral transduction followed by antibiotic selection if supported by the backbone (not specified for this item).

Reaction Conditions

Item-specific facts (from Product Data)

• None provided for experimental conditions.

General/literature guidance for typical workflows (non-item-specific)

• Transfection (plasmid delivery): Optimize DNA amount (e.g., 0.5–2 µg per well of a 6-well plate) and reagent ratio per manufacturer instructions of the transfection reagent; maintain cells at 60–80% confluence. Replace medium after 6–24 h as appropriate. Parameters are cell-line dependent.
• Lentiviral packaging: In third-generation systems, co-transfect the transfer plasmid with packaging plasmids in HEK293T cells, collect supernatant at 48–72 h, clarify, and optionally concentrate. Titer and transduce target cells at an MOI suitable for single-copy integration when desired. Follow BSL-2 practices and institutional approvals.
• Selection and verification: If the backbone provides a selection marker (not specified for this item), apply the appropriate antibiotic and validate editing by sequencing-based assays (e.g., amplicon NGS or Sanger followed by ICE/TIDE; literature).
• Cas nuclease provisioning: Supply Cas9 via co-transfection, stable expression cell lines, or RNP electroporation. Ensure the sgRNA scaffold is compatible with the chosen nuclease.

Note: The above are general literature practices and are not specific recommendations for this item. Refer to the CoA/Spec Sheet for any item-specific protocols.

Safety and Handling

Item-specific facts (from Product Data)

• GHS classification, pictograms, signal word, and H-statements: Not specified for this item; refer to SDS.
• Storage conditions: Store at −80°C.

General/literature guidance (non-item-specific)

• Biosafety: Plasmid DNA alone is typically handled at BSL-1. However, lentiviral transfer plasmids used for viral packaging and transduction should be handled under institutional BSL-2 practices when producing replication-incompetent lentivirus (literature and institutional guidelines). Follow your IBC-approved SOPs.
• PPE: Lab coat, safety glasses, and disposable nitrile gloves. Use nuclease-free technique and dedicated areas for DNA work to avoid contamination.
• Handling practices: Avoid repeated freeze–thaw; keep on ice during use. Use sterile, nuclease-free tips and tubes. Decontaminate surfaces with 10% bleach followed by ethanol; dispose of biological waste per local regulations.
• Incompatibilities: Avoid nucleases (DNase contamination), extreme pH (<6 or >9), and high temperatures. Avoid aerosol generation when preparing lentiviral particles.
• First aid (overview): In case of accidental exposure, rinse affected area with water. If eye contact occurs, rinse cautiously with water for several minutes. If ingested or inhaled in the context of aerosolized preparations, seek medical attention per SDS guidance. Always consult the product SDS for authoritative safety information.

Solvent Selection

This product is a plasmid DNA reagent, not a solvent. Classical solvent-selection criteria (polarity, dielectric constant, miscibility) do not apply.

General/literature handling notes (non-item-specific)

• Working medium: Plasmid DNA is typically handled in nuclease-free water or low-salt buffers (e.g., TE, pH ~7.5–8.0) to balance solubility and stability. High-ionic-strength buffers can affect downstream transfection or enzymatic steps.
• Additives: For long-term stability, EDTA (e.g., 1 mM in TE) can chelate Mg2+ to inhibit DNases (literature). For cell delivery, carrier solutions must be compatible with the chosen transfection/transduction method.
• For this item, any specific formulation or buffer requirements are not specified; consult the CoA/Spec Sheet.

Storage and Reconstitution

Item-specific facts (from Product Data)

• Storage conditions: Store at −80°C.
• Shipping conditions: Shipped on dry ice packs + cold packs.

Additional notes

• Reconstitution, working aliquots, and buffer formulation: Not specified for this item; refer to CoA/Spec Sheet.

General/literature best practices (non-item-specific)

• Upon receipt: Keep frozen; minimize time at ambient temperature. If supplied frozen, thaw on ice and mix gently by pipetting. Avoid vortexing to limit shearing.
• Aliquoting: Prepare single-use, nuclease-free aliquots to avoid repeated freeze–thaw cycles; store at −80°C for long-term stability.
• Contamination control: Use sterile, nuclease-free plasticware. Maintain a clean, DNase-free workspace.

For authoritative instructions on handling this specific lot/formulation, defer to the product’s CoA/Spec Sheet and SDS.

Structure and Identity

Item-specific facts (from Product Data)

• Product name: pLenti-CBL-sgRNA (SKU: P744457)
• Type: Plasmid DNA reagent (research use only)
• 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 / InChI / InChIKey: Not applicable to plasmid DNA; not specified for this item.

General/literature description (non-item-specific)

• Structural class: Circular double-stranded DNA (dsDNA) plasmid designed for CRISPR sgRNA expression in a lentiviral (pLenti) backbone (literature).
• Typical architecture (literature): An sgRNA expression cassette (often a U6 or H1 promoter driving a gRNA spacer fused to a tracrRNA scaffold), a lentiviral transfer backbone with long terminal repeats (LTRs), an origin of replication for E. coli propagation, and an antibiotic selection marker for bacterial maintenance. Specific features for this item are not provided.
• Target concept: “CBL-sgRNA” indicates the guide spacer is intended to target the CBL gene; the actual spacer sequence, promoter(s), selection marker(s), and backbone length are not specified for this item.
• 2D/functional description: Functionally, the plasmid encodes a small gRNA transcript that complexes with a CRISPR nuclease (e.g., Cas9 supplied separately) to direct site-specific DNA cleavage at the CBL locus (literature). No item-specific sequence map is provided here; consult CoA/Spec Sheet for definitive identity details.

Synthetic Utility

This section is framed for chemical building blocks; for a plasmid DNA reagent, “synthetic utility” translates to molecular cloning and genome-engineering utility.

General/literature utility (non-item-specific)

• Vector backbone: Serves as a chassis for cloning different sgRNA spacers targeting CBL or alternative genes. Common assembly methods include Golden Gate (Type IIS restriction) and Gibson Assembly.
• Modular design: Many pLenti sgRNA vectors support swapping promoters, selection markers, or reporters to suit experimental needs. Verify compatibility with the intended Cas nuclease (e.g., SpCas9 vs SaCas9) and any PAM constraints.
• Retrosynthetic planning (molecular biology): Design PCR amplicons with overlaps for seamless assembly; confirm the absence of internal Type IIS sites if using Golden Gate; include barcodes or unique cut sites for rapid screening.

Item-specific facts (from Product Data)

• Specific restriction sites, antibiotic resistance, promoter(s), and sgRNA scaffold details are not specified for this item; consult CoA/Spec Sheet for exact cloning features.

Target Specificity

Only information from Product Data may be listed here.

• Antigen/target: Not specified for this item. The product name suggests an sgRNA directed toward the CBL gene, but the spacer sequence, species, and any validated target sites are not provided in the Product Data.
• Clone/isotype/species reactivity: Not applicable; this is a plasmid reagent, not an antibody. No additional specificity data are provided in the Product Data.