NEW! TransIT-X2¢â Dynamic Delivery SystemA novel, polymeric system for efficient delivery of plasmid DNA and/or siRNA/miRNATransIT-X2 Dynamic Delivery SystemÀº non-liposomal, polymeric systemÀ» ÀÌ¿ëÇÏ¿© primary cellÀ» Æ÷ÇÔÇÑ ´Ù¾çÇÑ cell type¿¡ ¶Ù¾î³ È¿À²À» º¸ÀÔ´Ï´Ù.
TransIT-X2 Dynamic Delivery SystemÀº DNA¿Í siRNA/miRNAÀÇ °¢°¢ÀÇ transfection¿¡ ÀÌ¿ëÇÒ ¼ö ÀÖÀ¸¸ç, DNA¿Í siRNA/miRNA transfection °¡´ÉÇÕ´Ï´Ù.
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TransIT-X2 Dynamic Delivery SystemÀº °ÅÀÇ ¸ðµç cell type¿¡¼ transfectionÀÌ Lipofectamine2000 Á¦Ç°º¸´Ù ¿ì¼öÇÑ ¼º´ÉÀ» °¡Áö°í ÀÖ½À´Ï´Ù.
Figure 1. Visualization of High GFP Expression using TransIT-X2¢â Dynamic Delivery System. TransIT-X2¢â Dynamic Delivery System was used to transfect plasmid DNA encoding EGFP into A549, CHO-K1, HepG2, LNCaP, MDCK, PC12, primary human mammary epithelial cells (HMEC) and normal human dermal fibroblasts (NHDF). Transfections were performed in 35 mm MatTek dishes using 4-8 µl of TransIT-X2¢â to deliver 2 µg of DNA. Images (32X) were captured at 48 hours post-transfection using a Zeiss Axiovert S100 inverted fluorescence microscope.
* indicates primary cell types Figure 2. High GFP Transfection Efficiency in Multiple Cell Lines and Primary Cells using TransIT-X2¢â Dynamic Delivery System. TransIT-X2¢â Dynamic Delivery System was used to transfect plasmid DNA encoding EGFP into A549, CHO-K1, Hep G2, MDCK, LNCaP, PC-12, primary human mammary epithelial cells (HMEC) and normal human dermal fibroblasts (NHDF). Transfections were performed in 96-well plates using 0.2-0.4 µl of TransIT-X2¢â to deliver 0.1 µg of DNA (2:1, 3:1 or 4:1 reagent: DNA ratio). Triplicate wells were assayed 48 hours post-transfection using guava easyCyte¢â 5HT Flow Cytometer.
*indicates primary cell types Figure 3. Functional Co-delivery of Plasmid DNA and siRNA using the TransIT-X2¢â¢â Dynamic Delivery System. TransIT-X2¢â Dynamic Delivery System was used to transfect plasmid Cy¢ç5 labeled DNA encoding nuclear YFP and Cy¢ç3 labeled siRNA into HeLa cells. Transfection was performed in 6-well plates with Poly-L-Lysine (PLL) coated coverslips using 4 µl of TransIT-X2¢â to deliver 2 µg of DNA and 25 nM siRNA (2:1 reagent:DNA ratio). Actin cytoskeleton was stained using Alexa Fluor¢ç 350 Phalloidin. Images (63X) were captured at 24 hours post-transfection using a Nikon A1R confocal microscope. Image key: yellow (nuclear YFP), blue (Cy¢ç5 labeled DNA), red (Cy¢ç3 labeled siRNA), green (actin cytoskeleton).
Figure 4. TransIT-X2¢â Dynamic Delivery System Achieves Higher Knockdown than Lipofectamine¢ç 2000. TransIT-X2¢â Dynamic Delivery System and Lipofectamine¢ç 2000 Transfection Reagent were used to transfect siRNA targeting endogenous proteins – GAPDH and AHA1 or to deliver a non-targeting siRNA control in normal human dermal fibroblasts (NHDF). Cells were transfected in a 6-well plate using 4 µl of TransIT-X2¢â or 6 µl of Lipofectamine¢ç 2000 and 25 nM siRNA according to each manufacturer's protocol. The amount of GAPDH or AHA1 mRNA was measured relative to 18s rRNA levels using qRT-PCR and then normalized to the mRNA levels of the non-targeting control, 48 hours post-transfection. Error bars represent the standard deviation of triplicate wells.
Figure 5. Effective miRNA Delivery using TransIT-X2¢â Dynamic Delivery System Yields Decreased Levels of PTK9 mRNA. TransIT-X2¢â Dynamic Delivery System and Lipofectamine¢ç¢ç 2000 Transfection Reagent were used to transfect Pre-miR¢â hsa-miR-1 miRNA Precursor or mirVana¢â miRNA mimic, miR-1, both known to decrease PTK9 mRNA levels. A Pre-miR negative control was also transfected to assess baseline mRNA levels. T47D cells were transfected in a 12-well plate using 3 µl of TransIT-X2¢â or Lipofectamine¢ç 2000 and 50 nM miRNA according to each manufacturer's protocol. The amount of PTK9 mRNA was measured relative to 18s rRNA levels using qRT-PCR and then normalized to the mRNA levels of the negative control, 48 hours post-transfection. Error bars represent the standard deviation of triplicate wells.
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