I was wondering if anyone has tried transfecting human MSCs with an shRNA plasmid using the HUSH brand of plasmids from Origene along with their transfection reagent turbofectin 8.0. I am having limited success so far using this method and the plasmid has a GFP reporter to visualize the efficiency of transfection. So far I have only attempted to transfect with 1ug of plasmid DNA with a 1:3, 1:4, 1:7 and 1:10 ratio with turbufectin 8.0. I was wondering if more plasmid DNA is required or not. Also if someone has had better luck with another type of method I would love to hear about it. Thank you.
Hi Chris,
I have tried transfecting hMSCs (in house prepared & adipose derived) with just a GFP plasmid at different amounts and the efficiency is poor (~20-30%). I highly recommend not wasting your time/effort/money in attempting to transfect hMSCs. You will not get high efficiency.
I use lentivirus (made and titered in house) to transduce hMSCs and can achieve ~98% efficiency using an MOI of 5. You probably can find your shRNA of interest already packaged in a lentivirus from a company. If not, you can always make your own. All of the plasmids for making lentivirus are available through Addgene and are very inexpensive.
Good luck!
Tom
Hello Chris Delavan
For plasmid transfection into MSCs, I would highly recommend a product called Viromer Red, which is sold by Lipocalyx, and distributed by OriGene to US customers. Please see the data linked below.
Viromer Red stem cell transfection data: https://viromer-transfection.com/data-by-cell-type/stem-cell-transfection
OriGene Viromer free samples: https://www.origene.com/promotions/free-viromer-sample-for-you
Tom, yes I agree with you from what I have read in the literature that viral infection is the way to go when dealing with MSCs. My plasmid is retroviral instead of lentiviral so I am going to package it in-house with a retroviral packaging kit and attempt to transduce the plasmid that way.
Sonia Merritt that viromer red looks promising. What has been your experience using it to transfected hMSCs? What type of transfection efficiencies have you obtained with it and plasmids?
Chris Delavan my apologies for the slow response. Unfortunately, we do not have specific data from our company, however some of our customers have seen transfection efficiencies between 50-80% in hMSCs.
I will include a reference from someone who used the Viromer yellow product successfully. The Viromer red is thought to be more efficient than the yellow, however so far we only have a publication using Viromer yellow.Article Hypoxia Differentially Modulates the Genomic Stability of Cl...
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