Comparison of transfection efficiencies of a few in vitro DNA transfection reagents: GeneExpresso, LipoExpress, Lipofectamine 2000, and Expressfect

GeneExpresso and LipoExpress are from Lab Supply Mall
Lipofectamine 2000 is from Invitrogen.

Expressfect is from Denville Scientific, Inc. (http://www.denvillescientific.com), Expressfect is an OEM product (SoFast) from China (http://www.sunmabio.com, http://www.actgene.com/Convoy.htm).

Lipofectamine 2000	LipoExpress
GeneExpresso	Expressfect (SoFast)
Efficiency comparison of GeneExpresso, LipoExpress, Lipofectamine 2000, and Expressfect on Hela cells. A nuclear targeted GFP DNA was transfected with GeneExpresso, LipoExpress, Lipofectamine 2000, and Expressfect respectively per manufacturer’s protocols. The efficiency was checked 48 hours post transfection.
FACS Analysis

Conclusion:
Both GeneExpresso and LipoExpress have higher transfection efficiency than Lipofectamine 2000. Expressfect (OEM of SoFast) should not be used as transfection reagent.

GRGreen is a nucleic acid stain for detecting nucleic acids, e.g., double-stranded DNA, in agarose gel. It can be used for replacing mutagenic ethidium bromide (EB).   

Compared to EB which is a very strong mutagen, GRGreen caused fewer mutations than EB in the Ames test.

• Available at 10,000X in H₂O for better safety.
• Compatible with UV or blue light transilluminator and common gel documentation systems.
• Will not affect downstream experiments: compatible with all gel purification kits tested, will not inhibit ligation reaction etc.
• Compatible with Sodium Borate Electrophoresis Buffer: Run gel 2-3 times faster at higher voltage, resolve shaper bands in minutes, and less heat generation.
• Cut out DNA bands for subclonning under safer blue light: No mutations caused by EB and UV light.

Stain A	GRGreen
DarkReader Blue Light Transilluminator
Stain A	GRGreen
UV Transilluminator

 Fig. 2, GRGreen in water is more sensitive than Stain A in water.

1% agarose gel prepared with SeaKem LE Agarose (Lonza) and 1X GRGreen or 1X Stain A (from competitor A), run with 1X Lithium Borate Fast Electrophoresis Buffer at 100 V.

Left: 100 bp DNA ladder, 250ng, 125 ng, 62.5 ng and 31.3 ng, total ng DNA per lane.

Right: 1kb DNA ladder, 250ng, 125 ng, 62.5 ng and 31.3 ng, total ng DNA per lane.

TAE	TBE	Sodium Borate

Fig. 3, GRGreen is compatible with three common running buffers.

1% agarose gel prepared with SeaKem LE Agarose (Lonza) and 1X GRGreen, run with 1X  three Electrophoresis Buffers at 100 V.

Storage: Store at 4 ^oC.

Protocol:

1. Prepare 20 to 40 ml of agarose gel solution (concentration from 0.7~2.0%) with TAE, TBE or Borate Buffer in a 250 ml flask and mix it thoroughly. Place the flask in the microware, heat on high until the solution is completely clear and no small floating particles are visible (about 2~3 minutes).

2. After the gel solution cool to about 55 oC, add GRGreen to the solution to 1X final concentration. Swirl the flask gently to mix the solution and avoid forming bubbles.

3. Pour the gel solution into a gel tray until the comb teeth are immersed about 1/4~1/2 into the gel solution.

4. After the agarose gel has solidified you can perform electrophoresis using either 0.5 to 1X TAE, TBE or Borate Buffer.

5. Detect the bands using UV or blue light transilluminator.

FAQ

1, Should I wear gloves when using this dye?

   You should exercise common safe laboratory practices when using this reagent.

2, What blue light transilluminator should I used with GRGreen dye?

    DarkReader.

3, What filter should I use for blue light transilluminator?

    Filter from Clare Chemical.

 MSDS: GRGreen MSDS, Disposal Guide

Protocol

PolyExpress™ DNA In Vitro Tranfection Reagent is a biodegradable polymer based DNA transfection reagent. It is effective for transfecting of common   (HEK293, COS-7, NIH-3T3, HeLa, CHO) and  a broad ranges of hard-to-transfect mammalian cells. PolyExpress™ reagent is capable to immobilize DNA migration during electrophoresis at very low concentration and form polyplexes within a few minutes at room temperature. Due to its biodegrable feature,  the cationic polymer is rapidly degraded shortly after entering cells by endocytosis (Figure 1), therefore, it has much less cytotoxicity than other lipisome-based transfection reagent. PolyExpress™  (1.0 ml) is sufficient for 300 to 600 transfections in 24 well plates or 150 to 300 transfections in 6 well plates.

Figure 1. A Cartoon Showing Biodegradation of PolyExpress DNA Transfection Reagent After Endocytosis of Transfection Complex

Features

Bio-degradable after endocytosis

- Exceptional high titers of virus production

- Efficient for very long DNAs (>89 kb)

- Efficient for both single DNA transfection and multi DNA co-transfection

- High levels of recombinant protein production

- Simple and robust transfection procedure

- Very affordable

Storage Condition:  Store at 4 °C. If stored properly, the product is stable for 12 months or longer.

Broad Transfection Spectrum for Mammalian Cell Types

Examples Showing Transfection Efficiency of PolyExpress™ DNA In Vitro Transfection Reagent on Common Cell Lines

PolyExpress™	Lipofectamine Plus

Fig. 2, Transfection efficiency comparison of PolyExpress™ vs. lipofectamine Plus on Chinese Hamster Ovary (CHO) cells. HA tagged beta-tubulin cDNA was delivered into CHO cells with PolyExpress™ (left panel) and lipofectamine Plus (right panel) respectively. FITC conjugated antibody against HA tag was utilized to pick up HA-beta-tubulin (Green) while a DM1a antibody was used to detect endogenous alpha-tubulin followed by probing with rhodamine conjugated secondary antibody (Red). (Courtesy of Dr. Shang Yin, University of Texas at Houston Medical School).

PolyExpress™	Lipofectamine 2000

Fig. 3. A comparison showing transfection efficiency of PolyExpress™ reagent vs. a leading product, Lipofectamine 2000 on HEK293FT cells. HEK-293FT cells were transfected with GFP vector (pEGFP-N3) by PolyExpress™ (left panel) and Lipofectamine 2000 (right panel) respectively. The cells were visualized by Nikon Eclipse Fluorescence microscope 24 hours post transfection.

PolyExpress™	Lipofectamine 2000

Fig. 4. A comparison showing transfection efficiency of PolyExpress™ reagent vs. a leading product, Lipofectamine 2000 on HepG2 cells. HepG2 cells were transfected with GFP vector (pEGFP-N3) by PolyExpress™ (left panel) and Lipofectamine 2000 (right panel) respectively. The cells were visualized by Nikon Eclipse Fluorescence microscope 24 hours post transfection

PolyExpress™	Lipofectamine 2000

Fig. 5. A comparison showing transfection efficiency of PolyExpress™ reagent vs. a leading product, Lipofectamine 2000 on MDCK cells. MDCK cells are notoriously hard to transfect. With proprietary “Shaved Cell Transfection” protocol, PolyExpress™ (left panel) gives up to 70% GFP positive cells vs. Lipofectamine 2000 (right panel) around 5% efficiency. MDCK cells were transfected with GFP vector (pEGFP-N3) by PolyExpress™ (left panel) and Lipofectamine 2000 (right panel) respectively. The cells were visualized by Nikon Eclipse Fluorescence microscope 36 hours post transfection.

PolyExpress™	Fugene HD

Fig. 6. A comparison showing transfection efficiency of PolyExpress™ reagent vs. a leading product, Fugene HD on LNCap cells.  LNCap cells were transfected with GFP vector (pEGFP-N3) by PolyExpress™ (left panel) and Fugene HD (right panel) respectively. The cells were visualized by Nikon Eclipse Fluorescence microscope 24 hours post transfection

PolyExpress™ DIC	PolyExpress™ FITC

Fig. 7. Neuro2A cells transfected with pEGFP-C1 plasmid using PolyExpress™ In Vitro DNA Transfection Reagent. The Neuro2A cells were visualized by Nikon Eclipse Fluorescence microscope with DIC phase imaging (left) and FITC imaging (right) 24 hours post-transfection 

PolyExpress™	Lipofectamine 2000

Fig. 8. Comparison of cytotoxicity of PolyExpress ™ DNA In Vitro Transfection Reagent with L2K™ on primary murine skin fibroblast. The primary murine fibroblast was incubated with the indicated transfection reagents/pEGFP-C1 (DNA) complexes above for 4 hours in serum-free DMEM High Glucose medium followed by replacement of complete serum-containing medium. The cells were visualized by Nikon Eclipse Fluorescence microscope with DIC phase imaging 24 hours post transfection