PEG-TE-Lithium Acetate Solution (A+B+C, 8:1:1) - BioReagent

Cat. No.: P1528883

GRADE & PURITY BioReagent

Storage

Room temperature

Shipped In

Normal

Panoramica

This product is a key reagent for yeast transformation, consisting of Tris, EDTA, PEG3350, and lithium acetate in deionized water, with the TE-lithium acetate component sterilized by autoclaving. The three components (A:B:C) are used in an 8:1:1 ratio for the preparation of yeast competent cells and exogenous DNA transformation. Mechanism of Action • PEG3350 (polyethylene glycol, MW 3,350): In the presence of high concentrations of lithium acetate, PEG protects the yeast cell membrane from excessive damage, while promoting closer contact between plasmid DNA and the cell membrane, significantly enhancing transformation efficiency. Studies have shown that PEG3350 can achieve transformation efficiencies of up to 10⁵ cfu/μg DNA. PEG with a molecular weight of approximately 3,000 exhibits optimal transformation-promoting activity, and PEG3350 was selected based on this principle. • Lithium Acetate (LiAc): Induces a transient competent state in yeast cells, increasing membrane permeability and enabling the uptake of exogenous DNA. • TE Buffer (Tris-EDTA): Maintains pH stability, while EDTA chelates metal ions to protect DNA from nuclease degradation. Important Note The concentration of PEG is critical for transformation efficiency. Strict adherence to the recommended ratio is essential to ensure consistent and reproducible results.

Specifications

Specifiche e purezza

BioReagent

Condizioni di conservazione di stoccaggio

Room temperature

Spedito in

Normal

Grado

BioReagent

Product Line

Prodotti biologici

Application Protocols

• Item-specific note

  • No proprietary protocol was provided in the Product Data. The following is a concise, literature-style example protocol for yeast transformation using a PEG/TE/LiAc mixture. Adjust volumes and times to your system.

• Example protocol (literature)

  1. Grow yeast to OD600 ~0.4–0.8 (mid-log). Pellet and wash cells once with sterile water, then resuspend in LiAc-containing buffer.
  2. For each transformation, combine in a microtube: competent cells, 0.1–1 µg plasmid DNA, and 10–100 µg denatured salmon sperm carrier DNA.
  3. Add PEG-TE-LiAc solution to the specified final composition (product-specific concentrations Not specified for this item; refer to CoA/Spec Sheet). Vortex briefly to mix.
  4. Incubate at room temperature for ~30 min (or per your established method), then heat shock at ~42°C for 15–45 min.
  5. Pellet cells gently, remove supernatant, and resuspend in 0.2–1 mL non-selective medium. Recover with shaking for 0.5–3 h.
  6. Plate appropriate dilutions onto selective agar and incubate until colonies form.

• Tips

  • Ensure DNA is free of inhibitors (salts, phenol, ethanol). Keep PEG solution homogeneous; viscous mixtures may require thorough mixing.
  • Include negative controls (no DNA) and positive controls (well-characterized plasmid) to benchmark efficiency.

Biological Roles

• Item-specific note

  • This is a laboratory reagent mixture; not a biological metabolite. No organismal “role” is implied for the product itself.

• Component roles in workflow (literature)

  • PEG: acts as a macromolecular crowding agent, promoting DNA-cell proximity and membrane perturbation; can induce osmotic stress aiding uptake.
  • Lithium acetate: lithium ions increase cell wall permeability and competency in yeast; acetate provides buffering capacity within the system.
  • TE buffer (Tris-EDTA): Tris maintains pH near neutrality; EDTA chelates Mg2+/Mn2+, suppressing nuclease activity and stabilizing nucleic acids during handling.

• Context of use

  • Widely applied in Saccharomyces cerevisiae genetics, cloning, and genome engineering to introduce plasmids, cassettes, or CRISPR components (research use only).
  • Compatible with co-transformation strategies that rely on yeast homologous recombination machinery for in vivo assembly of DNA fragments.

• Limitations (literature)

  • Efficiency can vary by strain, plasmid size, DNA purity, and heat-shock conditions.
  • EDTA can inhibit downstream enzymatic steps if carried over at high levels; appropriate recovery and washing steps mitigate this.

Buffer Applications

• Applicability

  • Directly relevant: the reagent contains TE buffer and functions as an aqueous medium during transformation workflows.

• TE buffer context (literature)

  • Composition: Tris (commonly 10 mM) and EDTA (commonly 1 mM) at pH ~7.5–8.0 in standard laboratory practice; concentrations in this product are Not specified for this item; refer to CoA/Spec Sheet.
  • Function: maintains pH and protects DNA from nuclease degradation via EDTA chelation of divalent cations.

• Practical tips (literature)

  • If downstream steps are cation-dependent (e.g., transformation recovery, enzymatic reactions), dilute or remove EDTA carryover by washing cells after heat shock.
  • For sensitive applications, confirm the pH and ionic strength of the working mix; buffer capacity should support the brief heat-shock step without significant drift.
  • Sterile handling (filter or autoclave-compatible components prepared separately) reduces nuclease and bioburden risk.

• Non-applicable aspects

  • Not intended as a general electrophoresis or cell culture buffer by itself; its formulation prioritizes transformation efficiency rather than long-term cell maintenance.

Green Alternatives

• Environmental context (general)

  • The matrix is aqueous, avoiding volatile organic solvents. Components are of relatively low acute toxicity; however, EDTA is persistent in the environment and can mobilize metals, and PEG can contribute to chemical oxygen demand in wastewater.

• Alternatives and trade-offs (literature)

  • Electroporation: reduces chemical additives (no EDTA/PEG in the transformation mix) but increases electrical energy use and requires specialized equipment; cuvettes are often single-use plastic.
  • Spheroplasting with enzymes: can avoid EDTA but uses biological enzymes and osmotic stabilizers; generates different waste streams and may have higher cost/biological resource use.
  • Proprietary “greener” crowding agents: Some labs explore biodegradable polymers or lower-EDTA buffers; performance equivalence must be validated.

• Good practice with this reagent

  • Use only the volumes needed; keep solutions sterile to extend usable life and reduce waste.
  • Collect EDTA-containing wastes separately where required by local regulation; avoid discharge to aquatic systems.
  • Consider miniaturizing transformation reactions (micro-scale) to reduce consumables and buffer volumes without compromising outcomes.

Pharmaceutical Uses

• Not applicable for therapeutic or clinical use. This product is designated BioReagent grade and is for research use only (as stated in Product Data).
• General context (literature)
  • Components such as PEG may serve as excipients in pharmaceutical formulations in other contexts, and acetate salts are used as buffering agents. However, no pharmacopeial status, excipient grade, sterility assurance level, or GMP compliance is indicated for this item.
• For any application beyond research laboratory use, including manufacturing or diagnostic workflows, users must verify suitability, regulatory requirements, and quality attributes independently. No such claims are made for this product.

Physical Properties

• Item-specific properties

  • Appearance: Not specified for this item; refer to CoA/Spec Sheet.
  • pH, conductivity, osmolality, PEG concentration, LiAc molarity: Not specified for this item; refer to CoA/Spec Sheet.
  • Density, refractive index: Not specified for this item; refer to CoA/Spec Sheet.

• General properties of similar PEG/TE/LiAc mixtures (literature, typical)

  • Physical state: clear to slightly opalescent aqueous solution; viscosity increases with PEG content.
  • Miscibility: fully water-based; miscible with water; immiscible with nonpolar solvents.
  • pH range: governed by TE buffer, often near neutral to mildly basic (commonly ~7.5–8.0 in many protocols).
  • Ionic content: presence of Li+ and acetate; EDTA chelates trace divalent cations.
  • Temperature sensitivity: components are stable at ambient conditions; elevated temperatures can increase PEG viscosity and may accelerate microbial growth if not sterile.

• Notes

  • As a formulated mixture, classical single-compound descriptors (bp, mp, logP) are not applicable.
  • For exact specification values (PEG %, TE concentration, LiAc molarity, sterility), consult the product’s CoA/Spec Sheet.

Quality and Grades

• Item-specific quality

  • Grade: BioReagent (as provided in Product Data).

• What BioReagent typically means (general guidance)

  • Suitable for common molecular biology/biochemistry workflows (e.g., DNA handling, transformation protocols), emphasizing low levels of impurities that could inhibit enzymatic or cellular processes.
  • Not equivalent to clinical/diagnostic-grade or GMP materials; intended for research use only.

• Considerations for this mixture

  • PEG quality can influence viscosity and performance; EDTA/Tris purity impacts nuclease inhibition and pH stability; lithium acetate purity affects cellular compatibility. BioReagent grade generally aligns with these expectations.
  • UV background, nuclease-free status, sterility, endotoxin level, and bioburden: Not specified for this item; refer to CoA/Spec Sheet.

• Practical notes

  • If downstream applications are nuclease-sensitive, verify “DNase/RNase-free” or filtration status on the CoA.
  • For reproducibility, confirm batch-specific pH, PEG percentage, and Li+ molarity prior to critical work (consult CoA/Spec Sheet).

Reaction and Applications

• Item-specific scope

  • Manufacturer Applications: Not specified in Product Data beyond the product name. The following expands on common literature usage of PEG/TE/LiAc mixtures for research workflows.

• Literature applications

  • Yeast transformation (LiAc/PEG method): Facilitates uptake of plasmid or linear DNA by Saccharomyces cerevisiae and related yeasts via a combination of Li+-induced cell wall permeabilization and PEG-mediated macromolecular crowding.
  • DNA repair-based integration: Linear DNA with homology arms can integrate by homologous recombination in yeast following treatment with LiAc/PEG.
  • Co-transformation: Simultaneous introduction of multiple DNA fragments for in vivo assembly (e.g., gap repair cloning).

• Mechanistic notes (literature)

  • Li+ disrupts cell wall/membrane stability and enhances competency.
  • PEG promotes close apposition between DNA and cell surface, aiding uptake; TE protects nucleic acids (EDTA chelation reduces nuclease activity; Tris buffers pH).

• Practical guidance (literature)

  • Freshly prepare or vortex to homogenize prior to use; PEG solutions can separate slightly or form gradients.
  • Combine with denatured, sheared carrier DNA to enhance efficiency; heat-shock step (e.g., ~42°C) is typically applied.
  • Maintain clean, nuclease-minimized conditions to protect transforming DNA.

Reaction Conditions

• Item-specific parameters

  • No specific instructions were provided in the Product Data. The following are literature-based, general conditions for yeast chemical transformation using LiAc/PEG mixes; adapt to your strain and plasmid.

• Typical literature conditions (example framework)

  • Cell prep: Grow S. cerevisiae to mid-log phase; wash and resuspend in LiAc-containing buffer.
  • DNA: 0.1–1 µg plasmid DNA per reaction; include boiled, sheared salmon sperm DNA (carrier) at 10–100 µg to enhance efficiency.
  • Mix: Combine competent cells, DNA, carrier DNA, and PEG-TE-LiAc solution to reach the prescribed final PEG and Li+ concentrations (product-specific values Not specified for this item; refer to CoA/Spec Sheet).
  • Incubations: Pre-incubate on ice or at room temperature (10–30 min) depending on protocol; heat shock at ~42°C for 15–45 min (strain-dependent).
  • Recovery: Pellet, resuspend in non-selective medium, and outgrow 0.5–4 h before plating on selective medium.

• Expected outcomes (literature)

  • Transformation efficiencies vary widely (10^2–10^5 transformants/µg DNA), influenced by strain, DNA quality, and exact conditions.

• Notes

  • Maintain aseptic technique; verify the solution is homogeneous (PEG can stratify).

Safety and Handling

• Item-specific hazard data

  • GHS classification, signal word, pictograms, and H-statements: Not specified for this item; refer to SDS.

• General safety considerations (literature; not product-specific)

  • Potential hazards arise from components: lithium acetate (irritant), Tris buffers (low hazard, irritant), EDTA (irritant; chelator), and concentrated PEG (slippery spills; low acute toxicity).
  • Avoid contact with eyes/skin; prevent inhalation/ingestion. Ensure good laboratory hygiene.
  • PPE: lab coat, safety glasses, nitrile gloves. Consider face protection for large-volume handling.
  • Incompatibilities: strong oxidizers (rarely relevant in aqueous buffer work); avoid mixing with strong acids/bases that would disrupt buffer and release acetic acid.
  • First aid (general): rinse skin/eyes with water for ≥15 min if exposed; remove contaminated clothing; if inhaled (aerosol), move to fresh air; if ingested, rinse mouth — seek medical attention. Always follow the SDS.
  • Spills: PEG-containing solutions are slippery — mark area; absorb with inert material; wash residue with water.
  • Microbiological risk: Aqueous organic solutions can support microbial growth if contaminated; handle aseptically if sterility is required.

• Authoritative reference: Always consult the product SDS for definitive hazard classification, exposure limits, and emergency measures.

Solvent Selection

• Applicability

  • This product is an aqueous reagent mixture used as a transformation medium, not a general-purpose solvent for synthesis or extraction.

• General characteristics (literature)

  • Polarity: highly polar, water-based matrix.
  • Miscibility: fully miscible with water; not compatible with hydrophobic solvents.
  • Functional role: provides a crowding/precipitation environment (PEG) and ionic conditions (Li+) for facilitating nucleic acid uptake by cells, typically yeast.

• When to choose this formulation (context)

  • Prefer over simple saline or buffer alone when the LiAc/PEG mechanism is required to boost chemical transformation efficiency in yeast.
  • Alternatives: electroporation buffers for electrocompetent cells; spheroplasting media; proprietary high-efficiency transformation kits.

• Comparison snapshot (literature)

  • LiAc/PEG chemical transformation: inexpensive, scalable, no specialized equipment; moderate efficiency; tolerant to standard lab setups.
  • Electroporation: higher peak efficiencies with optimized strains and DNA; requires cuvettes/electroporator; sensitive to salt and contaminants.

• Practical tips

  • Maintain DNA in low-salt TE or water to avoid arcing in alternative methods and to control ionic strength when combining with this solution.
  • If mixing with other aqueous components (e.g., carrier DNA), confirm compatibility and final ionic strength.

Storage and Reconstitution

• Item-specific storage

  • Storage conditions: Room temperature (as provided in Product Data).
  • Shipped: Normal (as provided in Product Data).

• Stability and handling (general guidance)

  • Protect from microbial contamination; if sterility is required, work aseptically and consider aliquoting to minimize repeated opening.
  • PEG-containing aqueous solutions can exhibit viscosity changes with temperature; mix thoroughly before use to ensure homogeneity.
  • Do not freeze unless validated for your workflow; freeze–thaw cycles may cause component stratification or precipitation.

• Reconstitution

  • Not applicable — supplied as a ready-to-use solution. If concentration adjustments are needed, use sterile, nuclease-free water or compatible buffer; verify final composition does not deviate from your validated protocol.

• Shelf life and specs

  • Expiry, sterility status, and exact composition: Not specified for this item; refer to CoA/Spec Sheet.

• Research use only

  • As noted in Product Data: For research use only; not for diagnostic or therapeutic applications.

Structure and Identity

• Item-specific facts (from Product Data)

  • Product type: Ready-to-use mixture — PEG-TE-Lithium Acetate Solution (A+B+C, 8:1:1)
  • Grade: BioReagent
  • 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 — multi-component aqueous solution; Not specified for this item; refer to CoA/Spec Sheet.

• General description (literature)

  • Composition conceptually combines: polyethylene glycol (PEG), TE buffer (Tris•HCl/EDTA), and lithium acetate, commonly used for yeast DNA transformation.
  • “A+B+C, 8:1:1” denotes a volumetric blend of three stock components (literature conventionally: PEG solution + 10× TE + LiAc solution), yielding an aqueous transformation mix.
  • Functional entities: polymeric crowding agent (PEG), buffering/chelating system (Tris-EDTA), and lithium salt (Li+ acetate) that enhances cell wall permeability.

• Structural features (descriptive; literature)

  • PEG: linear poly(ethylene oxide) lacking discrete molecular formula (polymeric repeat –CH2–CH2–O–).
  • TE: buffering from Tris base (primary amine alcohol) around pH ~7.5–8.0; EDTA is a multidentate chelator.
  • Lithium acetate: small inorganic-organic salt providing Li+ and acetate anion in water.

Synthetic Utility

• This mixture is not a synthetic building block or a solvent for organic synthesis; therefore, conventional “synthetic utility” (e.g., named reactions, coupling chemistry) is not applicable.
• For users seeking chemical transformation utility in the biological sense, refer instead to Reaction & Applications and Application Protocols, which discuss its role in facilitating yeast transformation via LiAc/PEG methodology.

Target Specificity

• Not applicable. This product is not an antibody, probe, or targeted binder. It is a general reagent mixture for transformation workflows. No antigen/epitope, clone, isotype, or species reactivity information applies.

Domande frequenti

How should this product be stored?

Store at room temperature.