TY - JOUR
T1 - Neurobasal media facilitates increased specificity of siRNA-mediated knockdown in primary cerebellar cultures
AU - Gustafsson, Julie Ry
AU - Katsioudi, Georgia
AU - Issazadeh-Navikas, Shohreh
AU - Kornum, Birgitte Rahbek
N1 - Copyright © 2016. Published by Elsevier B.V.
PY - 2016/12/1
Y1 - 2016/12/1
N2 - Background Efficient and specific knockdown of proteins in post-mitotic cells such as differentiated neurons can be difficult to achieve. Further, special care must be taken to maintain the health of neurons in vitro. We wanted to achieve knockdown in primary cerebellar granule neurons, which can be effectively grown in Neurobasal™ media. New method We tested the efficiency of siRNA from the Accell range from Dharmacon™ when delivered in Neurobasal™ media in contrast to the recommended Accell Delivery media provided by the manufacturer. Results We observed a more specific knockdown of target in Neurobasal™ media, than in Accell Delivery media when using cerebellar granule neurons. Transfection efficiency and cell viability was comparable between the two media. Comparison with existing methods Delivery of siRNA in Neurobasal™ media facilitates increased specificity of the knockdown compared to delivery in Accell Delivery media. The off-target effect observed in Accell Delivery media was not a secondary biological response to downregulation of target, but rather a mixture of specific and non-specific off-target effects. Conclusions Specific knockdown of target can be achieved in primary cerebellar granule cells using Accell siRNAs in Neurobasal™ media.
AB - Background Efficient and specific knockdown of proteins in post-mitotic cells such as differentiated neurons can be difficult to achieve. Further, special care must be taken to maintain the health of neurons in vitro. We wanted to achieve knockdown in primary cerebellar granule neurons, which can be effectively grown in Neurobasal™ media. New method We tested the efficiency of siRNA from the Accell range from Dharmacon™ when delivered in Neurobasal™ media in contrast to the recommended Accell Delivery media provided by the manufacturer. Results We observed a more specific knockdown of target in Neurobasal™ media, than in Accell Delivery media when using cerebellar granule neurons. Transfection efficiency and cell viability was comparable between the two media. Comparison with existing methods Delivery of siRNA in Neurobasal™ media facilitates increased specificity of the knockdown compared to delivery in Accell Delivery media. The off-target effect observed in Accell Delivery media was not a secondary biological response to downregulation of target, but rather a mixture of specific and non-specific off-target effects. Conclusions Specific knockdown of target can be achieved in primary cerebellar granule cells using Accell siRNAs in Neurobasal™ media.
U2 - 10.1016/j.jneumeth.2016.10.001
DO - 10.1016/j.jneumeth.2016.10.001
M3 - Journal article
C2 - 27717866
SN - 0165-0270
VL - 274
SP - 116
EP - 124
JO - Journal of Neuroscience Methods
JF - Journal of Neuroscience Methods
ER -