□ 특징

● CRISPR/Cas9을 위한 sgRNA 및 mRNA, miRNA, siRNA등 여러 종류의 RNA Transfection에 최적

● 다양한 종류의 포유류 세포 (mammalian cell)에적용 가능

● 높은 효율의 유전자 도입과 매우 낮은 세포 독성

● Simple, serum-compatible protocol

● Transfection에 필요한 모든 구성품을 포함한 All-in-One system

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□ 제품설명

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Xfect™ RNA Transfection reagent는 다양한 종류의 mammalian cell (broad-range)에microRNA, siRNA, mRNA 또는 sgRNA와같은 여러 종류의 RNA를 효율적으로 도입 가능한Transfection 시약이다. 생분해성 나노입자 (biodegradablenanoparticles)로 구성된 Xfect™ Transfection reagnet는 일반적인 Transfection 시약에 비하여 낮은 세포독성및 높은 도입효율로 RNA Transfection이 가능하다. 특히 본 제품은 CRISPR/Cas9-mediated gene editing에 사용되는Cas9 coding mRNA 및 single guide RNA 도입에 효율적으로 사용이가능하며, Xfect™ Transfection reagent(Code 631317, 631318)과 함께 사용하면, DNA와 RNA co-transfection이 가능하다.그림 1. Xfect™ RNA Transfection 시약을 이용한 sgRNA Transfection 후,CD81 gene knock-out 결과 Panel A. sgRNA targeting the 5’ end of the antisensestrand of CD81 was synthesized using the Guide-it sgRNA In Vitro Transcription Kit. Panel B. An HT1080 cell line (2.0 x10⁵ cells) stably expressing Cas9 (HT1080-Cas9) was transfectedwith 50 pmol of sgRNA targeting CD81, either once or twice (lowergraph), using the Xfect RNA Transfection Reagent. Seven days later, cells wereimmunostained with a CD81 antibody (Ab) conjugated to an FITC fluorophore andanalyzed by flow cytometry. The percentage of cells that did not bind CD81 wascalculated. A control sample, comprised of HT1080-Cas9 cells, was analyzed byflow cytometry, either without (top, left graph) or with (top, right graph) theCD81 antibody. Both single and double transfection with sgRNA resulted in asubstantial increase in cells that did not bind CD81, indicating successfulCRISPR/Cas9-mediated knockout of CD81.그림 2. Primarycell 및 다양한 종류의 cell line에서의mRNA transfection 효과 HeLa cells (2.0 x10⁵), HEK 293 cells (1.5 x 10⁵), Human Dermal Fibroblasts(NHDF) cells (6.0 x 10⁴), Mesenchymal Stem Cells (MSCs) (5.0 x 10⁴),Jurkat Cells (3.0 x 10⁵), and KBM7 cells (3.0 x 10⁵) wereplated in 12-well plates and transfected with 1 μg of mRNA encoding GFP with 5μl of Xfect RNA Transfection Reagent. 20 hours later, cells were analyzed byflow cytometry and the % GFP-positive cells and the Mean Fluorescence Intensity(MFI) were determined.그림 3.Primary cell 및 cell line에서 siRNATransfection에 의한 Knock-down 효율HeLa cells (2.0 x 10⁵),Human Dermal Fibroblasts (NHDF) cells (6.0 x 10⁴), and MesenchymalStem Cells (MSCs) (5.0 x 10⁴)were plated in 12-well plates andtransfected with 50 pmol of siRNA against luciferase using Xfect RNATransfection Reagent.All three cell types were also transfected with 1 μg ofpCMV-Luc using the Xfect Transfection Reagent.Luciferase assays were performed 48 hourspost-transfection. For control samples, cells were transfected with pCMV-Lucbut without the siRNA.We observed a dramatic (>95%) decrease in luciferaseactivity in all the cells treated with siRNA.

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□ Applications

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□ 구성품

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□ 보존 - 20 ℃