pLenti-YTHDF2-sgRNA

Cat. No.: P751897

Spécifications

Product Name

pLenti-YTHDF2-sgRNA

Product Line

Produits biologiques

Stockage et expédition

Conditions de stockage de stockage

Store at -80°C

Expédié en

Dry ice packs + Cold packs

Stabilité et stockage

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

Application Protocols

Item-specific facts from Product Data

• Tested applications, recommended conditions, and positive controls: Not specified for this item; refer to CoA/Spec Sheet.

General/literature guidance (not item‑specific)

• sgRNA validation workflow: Sequence confirmation, small‑scale transfection with or without Cas9, editing assessment by TIDE/ICE or amplicon NGS, and phenotypic assay.
• Lentiviral use: Package with appropriate helper plasmids, titer, transduce target cells, and select/enrich if a selectable marker is present. Always follow BSL‑2 practices and institutional approvals.
• Controls: Non‑targeting sgRNA, safe‑harbor sgRNA, and rescue by sgRNA‑resistant cDNA where appropriate.

Biological Roles

Item-specific facts

• This product provides an sgRNA targeting YTHDF2; specific spacer sequence and species specificity are not specified for this item. Refer to CoA/Spec Sheet for the exact target sequence and genome build.

Literature context on the target (YTHDF2)

• YTHDF2 is an m6A reader protein that recognizes N6‑methyladenosine‑modified mRNAs and promotes their decay or redistribution, modulating mRNA stability and gene expression programs.
• Functional roles: Regulation of RNA turnover, stress responses, and transcriptome remodeling; often studied to dissect epitranscriptomic control mechanisms.
• Using sgRNA reagents: Disrupting YTHDF2 via CRISPR/Cas9 enables loss‑of‑function studies to attribute phenotypes to m6A‑dependent decay pathways and to map downstream transcript changes (e.g., by RNA‑seq or RIP‑seq; literature).

Caveats (general)

• Gene family/paralog considerations: YTH domain family includes YTHDF1/3; functional redundancy/compensation may occur—consider multiplex sgRNAs if appropriate.
• Off‑target effects: Use multiple independent sgRNAs and rescue experiments to validate on‑target causality (literature best practice).

No clinical/therapeutic claims are made. This product is for research use only.

Buffer Applications

Applicability

• Classical buffer systems for biochemical assays are not the primary focus for a plasmid reagent. However, storage and handling buffers are relevant.

General/literature guidance

• TE buffer (10 mM Tris‑HCl, 1 mM EDTA, pH 7.5–8.0): Common for long‑term plasmid stability; EDTA chelates Mg2+ to inhibit nucleases. May need dilution or buffer exchange before transfection/packaging.
• Nuclease‑free water: Optimal for immediate enzymatic use or transfection to avoid EDTA interference; store aliquots at −80 °C to limit degradation.
• Low‑EDTA Tris (10 mM, pH 7.5–8.0): Compromise between stability and enzymatic compatibility.

Practical tips

• For electroporation, use low‑ionic‑strength buffers to prevent arcing.
• If high salt is present (not specified for this item), perform ethanol precipitation or bead‑based cleanup prior to sensitive enzymology.

Item-specific note

• Exact buffer composition for this product is not specified; refer to CoA/Spec Sheet.

Green Alternatives

Applicability

• Green chemistry substitutions are less relevant for a nucleic acid reagent itself; however, associated workflows can adopt greener practices.

Greener practices for DNA work (literature guidance)

• Safer nucleic acid stains: Replace ethidium bromide with SYBR Safe or GelRed for gel electrophoresis.
• Solvent minimization: Prefer aqueous‑based cleanup (spin columns, magnetic beads) over phenol–chloroform extraction when possible.
• Reduced plastic waste: Use reusable gel trays, consolidate preparations, and adopt mini‑prep formats with lower plastic consumption.
• Energy: Store aliquots efficiently to minimize door‑open time at −80 °C; consider −70 °C if validated for your workflow.

Comparison snapshot (general)

• Phenol–chloroform vs. silica/magnetic cleanup: Silica/beads avoid halogenated solvents, reduce hazardous waste, and are typically faster, with comparable purity for most applications.

Note

• No alternative “green chemical” replaces the plasmid itself; improvements focus on ancillary reagents and procedures.

Pharmaceutical Uses

Applicability

• This is a research‑grade plasmid/sgRNA reagent. It is not produced under GMP and is not intended for human or veterinary therapeutic, diagnostic, or clinical use.

General/literature context

• Plasmid DNA can serve as a starting point in discovery workflows (e.g., target validation, mechanism studies) that precede translational development.
• For any clinical or manufacturing application, GMP-grade materials with full documentation, release testing, and validated processes would be required. This item does not include such claims or documentation.

Item-specific statement

• No pharmacopeial status or excipient role is specified for this item. For research use only.

Physical Properties

Item-specific facts (from Product Data)

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Concentration, solvent/buffer: Not specified for this item; refer to CoA/Spec Sheet.

General/literature values for plasmid DNA (for context; not item specifications)

• Physical state: Aqueous DNA solution or lyophilized solid depending on formulation.
• UV characteristics: dsDNA exhibits A260 with 1 A260 ≈ 50 µg/mL; typical purity A260/A280 ~1.8–2.0 (literature).
• Solubility: Highly water-soluble; insoluble in nonpolar organic solvents.
• pH stability: Stable near neutral pH; prolonged exposure to pH <6 or >9 accelerates depurination/strand breakage (literature).
• Temperature stability: Long-term storage at −80 °C recommended to minimize nuclease activity and hydrolysis (literature).
• Viscosity: Increases with concentration and supercoiled fraction (literature).

Not applicable measures for nucleic acids

• Boiling/melting point, density, logP, refractive index: Not applicable/reported as bulk macromolecule properties rather than fixed constants.

Practical notes

• Thaw on ice; mix gently to avoid shear. Use nuclease‑free, low‑EDTA buffers when downstream enzymology is intended. For accurate quantification, use A260 or fluorescent dsDNA assays (e.g., PicoGreen) with appropriate standards (literature guidance).

Quality and Grades

Item-specific facts (from Product Data)

• Grade/Purity: Not specified for this item; refer to CoA/Spec Sheet.
• Sequence/map verification: Not specified for this item; refer to CoA/Spec Sheet.

How to interpret quality for plasmid/sgRNA reagents (general guidance)

• Sequence identity: Verified by Sanger or NGS of the sgRNA spacer and key backbone regions (promoters, resistance markers). For CRISPR work, single‑base accuracy of the 20‑nt spacer is critical.
• Supercoiled fraction: High supercoiled content improves transfection and packaging efficiency; reported by analytical agarose or capillary electrophoresis.
• Endotoxin level: Endotoxin‑reduced or transfection‑grade DNA (<10 EU/µg typical for many applications) enhances cell viability; exact limit is not specified for this item.
• Residuals: Protein, RNA, salt carryover can impede enzymatic steps; OD260/230 provides a quick check (literature).
• Stabilizers/buffer: Presence of EDTA protects from nucleases but may inhibit downstream enzymology; exact buffer for this item is not specified.

Documentation

• For definitive specifications (concentration, buffer, resistance marker, plasmid length, sequence verification), consult the item’s CoA/Spec Sheet. Do not assume antibiotic marker or promoter without documentation.

Reaction and Applications

Item-specific facts (from Product Data)

• Manufacturer Applications: Not specified beyond the product name. General expansion provided below is literature‑based guidance and not item‑specific performance data.

Literature/general applications for pLenti‑sgRNA constructs

• CRISPR gene perturbation: The sgRNA targeting YTHDF2 can be used with a compatible Cas9 (stable line or co‑delivery) to induce double‑strand breaks and frameshift mutations via NHEJ, enabling knockout studies.
• Lentiviral workflows: When cloned in a lentiviral backbone, the sgRNA cassette can be packaged into pseudotyped particles for transduction of dividing and non‑dividing cells (performed under BSL‑2 with appropriate controls).
• Functional genomics: Use in pooled or arrayed screening to interrogate YTHDF2 function or synthetic interactions.
• Cloning and validation: Typical steps include sequence confirmation of the spacer, test transfection, cutting efficiency assays (e.g., T7E1, ICE/TIDE analysis), and downstream phenotyping.

Practical considerations (literature)

• Verify spacer: Confirm 20‑nt spacer and PAM context match the target genome version; assess predicted off‑targets with CRISPR design tools.
• Delivery: If Cas9 is not encoded in this vector, ensure cells express Cas9 (stable line, co‑transfection, or co‑transduction).
• Controls: Include non‑targeting and safe‑harbor targeting controls; consider multiple independent sgRNAs per gene.
• Antibiotic selection and markers: Not specified for this item—confirm before designing selection schemes.

Reaction Conditions

Applicability

• No chemical reaction conditions apply. For this reagent, “conditions” concern molecular biology workflows. The following are literature‑based guidelines, not item‑specific specs.

Typical workflows (literature)

• Transfection of plasmid DNA: Many cell lines at 37 °C, 5% CO2; reagent choice (lipid/PEI/electroporation) depends on cell type. DNA quality (supercoiled, low endotoxin) improves efficiency.
• Lentiviral packaging: Co‑transfect HEK293T or similar producer cells with the sgRNA vector and packaging/envelope plasmids; collect supernatants 48–72 h post‑transfection; filter and transduce targets under BSL‑2.
• Selection and validation: Apply antibiotic selection if encoded by the backbone (not specified for this item); validate editing by PCR and Sanger/NGS. Functional assays follow.

Operating parameters (examples; literature)

• DNA amounts: 0.5–2 µg per well of a 6‑well plate for transfection; scale as needed.
• MOI for transduction: Adjust to achieve desired single‑copy rates for clonal editing; include polybrene only if compatible with cells.

Important

• Backbone features (antibiotic marker, promoters, Cas9 presence/absence) are not specified for this item—verify documentation before setting conditions.

Safety and Handling

Item-specific facts (from Product Data)

• GHS classification, pictograms, signal word, H‑statements: Not specified for this item; refer to SDS.
• Storage: Store at −80 °C. Shipments on dry ice + cold packs.
• Research Use: For research use only.

General/literature safety guidance for plasmid DNA and lentiviral backbones

• Biosafety: Plasmid DNA itself is non-infectious; however, constructs bearing lentiviral elements are typically handled under BSL‑2 when used for packaging into viral particles. Do not generate replication‑competent virus; follow institutional biosafety approvals (literature/biosafety guidance).
• PPE: Lab coat, gloves, eye protection. Work in a clean area; use nuclease‑free technique to prevent contamination.
• Incompatibilities: Avoid nucleases (RNase/DNase) and microbial contamination. Do not mix with strong oxidants or extremes of pH that may degrade DNA.
• Aerosol minimization: Open tubes carefully; use filtered tips.
• First aid (general): If contact with eyes/skin, rinse with water. If ingestion/inhalation suspected, seek medical attention. Provide SDS to healthcare personnel.
• Spill/cleanup: Absorb aqueous spills with paper towels; decontaminate surfaces with 10% bleach followed by 70% ethanol (literature lab practice). Dispose according to local regulations and institutional biosafety procedures.

Always consult the product‑specific SDS and your institutional biosafety office for authoritative requirements.

Solvent Selection

Applicability

• As a plasmid DNA reagent, classical organic solvent selection is not applicable. The relevant “solvent” choice is the aqueous buffer for handling and storage.

General/literature guidance for plasmid DNA buffers

• Common solvents: Nuclease‑free water, TE buffer (10 mM Tris‑HCl, 1 mM EDTA, pH 7.5–8.0), or low‑EDTA Tris for enzymatic applications.
• Compatibility: DNA is fully miscible in water and aqueous buffers; avoid alcohols/strong chaotropes outside of purification contexts.
• When to choose which:
  • Nuclease‑free water: Ideal for immediate transfection or enzymatic reactions; maintain on ice and aliquot to minimize freeze–thaw.
  • TE buffer: Better long‑term stability due to EDTA chelation of Mg2+; may require dilution or desalting before transfection/packaging.
  • PBS: Acceptable for some cell delivery protocols; verify compatibility with reagent/vendor recommendations.

Practical tips

• Use low‑salt buffers for electroporation to minimize arcing.
• Filter tips and sterile technique reduce bioburden. Store in DNase‑free tubes and avoid repeated freeze–thaw cycles.

Storage and Reconstitution

Item-specific facts (from Product Data)

• Storage: Store at −80 °C.
• Shipping: Dry ice packs + Cold packs.
• Reconstitution and buffer: Not specified for this item; refer to CoA/Spec Sheet.

General/literature guidance

• Upon receipt: Keep frozen; minimize time at ambient temperature. Record lot information and, if provided, concentration and buffer.
• Aliquoting: Thaw on ice, mix gently, and aliquot into nuclease‑free tubes to avoid repeated freeze–thaw cycles.
• Short‑term storage: −20 °C (days to weeks) may be acceptable; for months to long‑term, retain at −80 °C.
• Buffer choices: Store in TE (10 mM Tris‑HCl, 1 mM EDTA, pH 7.5–8.0) for stability; switch to nuclease‑free water before sensitive enzymology or transfection.
• Avoid: Multiple freeze–thaw cycles, vigorous vortexing (can shear DNA), and nuclease contamination. Use sterile, filtered tips and DNase‑free plastics.

Note

• If supplied lyophilized or at a defined concentration, follow the exact reconstitution instructions on the CoA/Spec Sheet for solvent, target concentration, and mixing method.

Structure and Identity

Item-specific facts (from Product Data)

• Product: pLenti-YTHDF2-sgRNA (SKU: P751897)
• Category: Research reagent; plasmid/viral vector carrying an sgRNA targeting YTHDF2
• Storage: Store at -80°C
• Shipping: Dry ice packs + Cold packs
• 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 / CAS / CID: Not applicable to plasmid DNA constructs; not specified for this item.

General/literature description of structure

• Construct class: Lentiviral pLenti backbone bearing a single guide RNA (sgRNA) expression cassette targeting the human YTHDF2 locus (literature/typical for such naming).
• Functional elements usually present in pLenti-sgRNA vectors (literature): 5′/3′ LTRs, ψ packaging signal, antibiotic selection marker, origin(s) of replication, and a Pol III promoter (e.g., U6 or H1) driving the sgRNA scaffold. Exact backbone, selection marker, and promoters are not specified for this item.
• 2D “structure” description: Circular double‑stranded DNA plasmid; features include non-coding viral regulatory sequences and an sgRNA cassette; no small‑molecule functional groups.

Important note

• Exact plasmid map, length (bp), antibiotic resistance, promoters, and sgRNA spacer sequence are not specified for this item; refer to CoA/Spec Sheet or sequence file to confirm.

Synthetic Utility

Scope

• Organic small‑molecule synthetic utility does not apply. Instead, consider utility in synthetic biology and genome engineering.

Literature/general utility for pLenti‑sgRNA constructs

• Modular cloning: sgRNA spacer can be re‑cloned via BsmBI/BbsI‑based Golden Gate or Gibson Assembly into compatible backbones.
• Combinatorial perturbation: Facilitates pooled libraries or dual‑sgRNA strategies to study genetic interactions or large deletions (with paired cut sites).
• Retrosynthetic planning (synthetic biology): Backbone features (promoter, selection marker, origin) define compatibility with hosts and co‑vectors; careful planning prevents incompatibilities (e.g., origin clash in co‑transformations).
• Downstream edits: When co‑delivered with HDR donors, can direct precise edits; donor design includes homology arms and PAM disruption to prevent re‑cutting (literature).

Item-specific unknowns

• Exact cloning sites, antibiotic marker, and promoter(s) are not specified for this item; consult CoA/Spec Sheet to confirm before designing cloning strategies.

Target Specificity

Item-specific facts from Product Data

• Target, epitope/sequence, species reactivity, clone/isotype: Not specified for this item; refer to CoA/Spec Sheet.

Context

• The product name implies an sgRNA directed at YTHDF2, but the exact spacer sequence, PAM, and genome build are not provided here. Sequence‑level specificity and off‑target profile must be derived from the final validated spacer sequence provided in product documentation (if available).

Foire aux questions

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-YTHDF2-sgRNA); cross-reactivity in untested species may exist but is not guaranteed. Contact technical support for specific species inquiries.