Polyethylenimine Linear(PEI) MW 25000 _40815ES

Linearized Polyethyleneimine PEI 25000 Transfection Reagent is a highly charged cationic polymer that easily binds negatively charged nucleic acid molecules, forms a complex, and allows the complex to enter the cell. The transfection reagent is a transient transfection reagent with low cytotoxicity, high transfection efficiency, and high gene expression efficiency in cells such as HEK293 and CHO. It has been verified that the linear PEI transfection reagent is widely applicable to a variety of cell lines including HEK-293, HEK293T, CHO-K1, COS-1, COS-7, NIH/3T3, Sf9, HepG2 and Hela cells, etc. The reagent is compatible with serum-containing media and can efficiently introduce nucleic acids into cells.

Specification

 

Shipping and Storage

The product is shipped at room temperature and the powder can be stored at 2-8ºC for two years. The stock solution is stored at 2-8 ºC for 3 months.

Cautions


1) After the prepared PEI solution is taken out from -20 ºC and thawed, it can be stored in a 4 ºC refrigerator and must not be re-frozen.

2) For most cells, 3.0 μL of PEI transfection reagent per 1 μg of DNA can achieve high transfection efficiency. You can also try to use 1.5~4 μL volume of linear PEI transfection reagent per 1 μg DNA for optimization.

3) For your safety and health, please wear a lab coat, disposable gloves and a fume hood for operation.

4) This product is only for scientific research purposes, not for human use.

 

Stock solution Preparation (1 mg/mL)


1. Materials

PEI 25000, Milli-Q® water/water for injection (WFI) or similar biological grade water, 12 mol/L hydrochloric acid (HCl), 10 mol/L sodium hydroxide (NaOH), disposable 0.1~0.2 μm PES vacuum sterile filter, sterile HDPE or polypropylene storage bottle.

2. Prepare stock solution (1 mg/mL)

1) In a 1 L glass beaker, add 1 g of PEI 25000 powder into 900 mL of Milli-Q® ultrapure water or other equivalent biological water, and stir evenly on a magnetic stirrer to produce a small vortex.

2) Add hydrochloric acid (12 mol/L) dropwise while stirring to adjust the pH until pH<2.0.

3) Cover the top of the beaker and stir for 3 hours until completely dissolved; keep the pH < 2.0 throughout the process.

【Note】: There may be some small fibrous particles that cannot be dissolved, which is a normal phenomenon.

4) Add NaOH (10 mol/L) dropwise while stirring to adjust the pH until it reaches 6.9 ~7.1.

5) Transfer the solution into a graduated cylinder, and add water to make up to 1 L.

6) Filter and sterilize with a disposable 0.1-0.2 µm PES vacuum filter to obtain a 1 mg/mL stock solution.

7) Aliquot as needed and store at -20 °C, stable for 1 year.

【Note】: After re-thawing the storage solution, it can be stored at 4 °C and is stable for 2 weeks, but it must not be re-frozen

 

 

Transfection procedure (taking 6-well plate as an example)


1. Cell inoculation: In order to improve the transfection efficiency, it is recommended to inoculate the cells one day before transfection, and the cell density at the time of transfection is preferably 70%~80%.

2. Prepare the DNA-PEI complex: Prepare the DNA-PEI nucleic acid-transfection reagent complex according to the following system:

1) For each well of cells, dilute 2 μg of target DNA with 100 μL of serum-free medium, and mix thoroughly to form a DNA dilution.

[Note]: Opti-MEM or ddH2O is recommended for serum-free diluent

2) Immediately add 5 μL of PEI 25000 transfection reagent to 100 μL of DNA diluent, vortex for 10 seconds, and mix well.

3) Incubate at room temperature for 10-25 min to form DNA-PEI cationic nucleic acid transfection reagent complex.

3. Transfected cells:

1) During complex formation, remove the cell growth medium and add 2 mL of fresh pre-warmed complete medium to each well.

2) Add 100 μL DNA-PEI nucleic acid-PEI complex directly to the cells, shake the culture plate, and mix gently.

3) Cultivate in a 37°C, 5% CO2 incubator, and the expression of the transgene can be detected as soon as 7 hours after transfection. Please determine the suitable testing time by yourself.

4. Steady rotation screening (optional)

24 h after transfection, the cells were subcultured into fresh growth medium (dilute the cells more than 10 times), and incubated overnight at 37 °C in a 5% CO2 incubator. The next day, a screening drug matching the transfection resistance gene was added. Drug-resistant clones can be screened in about 1 to 2 weeks. During this period, the growth medium containing the screening drugs needs to be replaced frequently.

Transfection dosage for different cell culture vessels (for reference only):

Culture vessel	Surf. area per well*(cm2)	DNA(μg)	Transfection reagent(μL)	Vol. of dilution medium **(μL)	Vol. of plating medium
96-well	0.3	0.1	0.1	10	100μL
48-well	0.7	0.2	0.3	20	200μL
24-well	1.9	0.5	1	50	500μL
12-well	3.8	1	2	50	1mL
6-well	10	2	4	100	2mL
Flask  25cm²	21	4	8	200	4 mL
Flask  75cm²	58	10	20	500	10 mL