Transcript Assay result for cell line transfection

Requirement for a new transfection assay for RNAi
 Each cell has unique transfection efficacy.
For successful RNAi, good transfection method is essential.
One easy way is testing with Fluor-conjugated siRNA. As shown in below, the method
is for only transfection but not for RNAi.
Another method is testing gene silencing after transfection using qPCR, which
demands for cost and labor.
A fast and easy transfection assay for RNAi is required.
Fig. 1 Before (left) and after (right) transfection of CEM cell
with FITC-conjugated Vimentin siRNA.
Fig. 2 Total RNA sample was prepared after the transfection
and used for qPCR for test the silencing of target gene.
Unlike HeLa, there was no sign of gene silencing even
thought indication of good transfection by the FITCconjugated siRNA.
Transfection control siRNA
 The easiest and fastest way for siRNA transfection test
 Just add to the cell, wait for 48 hours, and count the viable cell
 The cell toxicity and level of the gene silencing is proportional
The target gene of the used siRNA is essential for cell viability
For human, mouse, and rat cells
Materials included
• Transfection control siRNA (2 nmole)
• Scramble siRNA (2 nmole)
• siRNA dissolving buffer
Transfection control siRNA
Cell viability of
MCF7 in 48 hour
Scramble siRNA 1 nM
control siRNA 1 nM
Gene silencing
in 24 hour
*All transfection assay was done by a transfection reagent G-Fectin.
The cell toxicity and level of the gene silencing is
proportional
MCF7
*All transfection assay was done by a transfection reagent G-Fectin.
The cytotoxic effect of Transfection control siRNA in
mouse and rat cells
*All transfection assay was done by a transfection reagent G-Fectin.
G-Fectin
1. A novel transfection reagent developed for RNAi
2. Whole transfection in 10 min on the same day. The
fastest and easiest method for transfection.
3. Your gene silencing in 24 hours
4. Lower toxicity
5. Good transfection efficacy
6. Compatible to siRNA. shRNA, and miRNA
7. 0.5ml (for 250 X transfection of 24 well plate)
2 ul of G-Fectin
siRNA
Add G-Fectin &
siRNA to 50 ul of
PBS in tube
10 min
transfection
protocol
with G-Fectin
Incubate 10 min at RT
Add the mixture
to plate directly
Detached the cell and add 450 ul
of media (5 x 104 cells/ well) to
24 well plate
No cell splitting is required in
previous day
Incubate 24 hrs
qPCR or Western
G-Fectin has good Transfection efficacy
Remaining Vimentin %
120
100
80
HeLa
60
OCUG
SNU368
40
Hep3B
MDA-MB-231
20
0
lipo 2000
G-factin
product 1
product 2
product 3
product 4
transfection reagent
Transfection assay using Vimentin siRNA final 10 nM
Tested cell line list
The Transfection efficacy was tested using the Transfection Control
siRNA and G-Fectin.
• Excellent transfection efficacy with 2ul of G-Fectin
HeLa, CHO, KB, HT1080, 3T3-L1, COS-1, Caco-2, SKOV3, MDAMB-468, HEP3B, siHa, MIA-PACA2, PANC1, PLC/PRF/5, MCF7,
PC3, C4I, J82, MRC5, HaCat
• Good transfection efficacy with 4 ul of G-Fectin
SNU-638, SNU-368, SNU-475, Noz, OVCA429, T98G,
• No transfection
Jurkat, CEM
G-Fectin has low cytotoxicity
All described cells were spitted in 30 % confluency in 36 hour ahead of viable cell
counting with treatment of lipo-plexed scramble RNA in final 10 nM.
G-Fectin Kit
 G-Fectin + Transfection Control siRNA
The easiest and fastest way for siRNA
transfection test
 Includes all reagents required for
transfection and transfection efficacy test
Can use to human, mouse, and rat
Materials included
• Transfection control siRNA (2 nmole)
• Scramble control siRNA (2 nmole)
• siRNA dissolving buffer
• G-Fectin 0.5ml (for 500 X transfection to 24 well)