What is the best transfection reagent?

Under the tested conditions, ViaFect™ Reagent offered the best combination of transfection efficiency and low toxicity for most cell lines, making it an ideal choice when beginning transfection experiments with a new cell line.

What is Lipofectamine reagent?

Lipofectamine 2000 Reagent is a cationic lipid-based transfection reagent designed to give extremely high transfection efficiencies and result in equally high protein expression levels. It has also been optimized to work for many mammalian cell lines.

What is transfection by chemical method?

The most commonly used method in clinical trials is the biological method where transfected nucleic acids are delivered to cells by viruses. The chemical transfection methods are techniques that catalyze DNA cross-membrane transport through the use of Ca2+phosphate, polycations or dendrimers.

How does calcium phosphate transfection work?

The calcium phosphate transfection method for introducing DNA into mammalian cells is based on forming a calcium phosphate-DNA precipitate. Calcium phosphate facilitates the binding of the DNA to the cell surface. DNA then enters the cell by endocytosis.

What does P3000 reagent do?

P3000 reagent: This reagent will be added to the diluted DNA solution, and its function is to help nucleic acids entering both the cell and the nucleus.

What is Lipofectamine made of?

Lipofectamine consists of a 3:1 mixture of DOSPA (2,3‐dioleoyloxy‐N‐ [2(sperminecarboxamido)ethyl]‐N,N‐dimethyl‐1‐propaniminium trifluoroacetate) and DOPE, which complexes with negatively charged nucleic acid molecules to allow them to overcome the electrostatic repulsion of the cell membrane.

What is transfection ratio?

The optimal ratio of transfection reagent to DNA is highly cell type-dependent. As a starting point, the amount of transfection reagent should be varied while keeping a constant plasmid DNA concentration (for example, 1:1, 3:1 and 5:1 ratios of volume to mass).

Why is calcium chloride used in transformation?

The addition of calcium chloride to a cell suspension promotes the binding of plasmid DNA to lipopolysaccharides (LPS). Positively charged calcium ions attract both the negatively charged DNA backbone and the negatively charged groups in the LPS inner core.

What is cacl2 transfection?

The procedure is routinely used to transfect a wide variety of cell types for transient expression or for producing stable transformants. The DNA is mixed directly with a concentrated solution of CaCl2, which is then added dropwise to a phosphate buffer to form a fine precipitate.

How do you create a stable transfection?

Stable cell line generation protocol

1. Generate a kill curve to determine the optimal selection antibiotic concentration.
2. Transfect cells with desired plasmid construct(s)
3. Select and expand stable polyclonal colonies.
4. Identify single clones by limited dilution and expansion.
5. Transfer clones and assess expression.

What is Lipofectamine P3000 reagent?

Lipofectamine™ 3000 reagent is designed to efficiently transfect difficult-to-transfect cells, yielding superior transfection performance across the broadest array of cell types. Successfully transfected cell types include: Fibroblast (3T3, COS-7) Myoblast (C2C12, L6 CRL-1458) Hepatocyte (HepG2, HuH7)

What does Lipofectamine 3000 do?

Lipofectamine™ 3000 reagent is suitable for novel genome editing applications. It increases the likelihood of successful cleavage and recombination with TALENs or CRISPRs through highly efficient transfection, ultimately maximizing the efficiency of genetic modifications and simplifying the downstream processes.

What is the difference between lipofectamine 2000 and 3000?

Lipofectamine 3000 reagent yields higher transfection efficiencies than Lipofectamine 2000 reagent when tested in a variety of cell lines.