Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

FOXA1 siRNA knock-down

SB Sylvan C. Baca
DT David Y. Takeda
JS Ji-Heui Seo
JH Justin Hwang
SK Sheng Yu Ku
RA Rand Arafeh
TA Taylor Arnoff
SA Supreet Agarwal
CB Connor Bell
EO Edward O’Connor
XQ Xintao Qiu
SA Sarah Abou Alaiwi
RC Rosario I. Corona
MF Marcos A. S. Fonseca
CG Claudia Giambartolomei
PC Paloma Cejas
KL Klothilda Lim
MH Monica He
AS Anjali Sheahan
AN Amin Nassar
JB Jacob E. Berchuck
LB Lisha Brown
HN Holly M. Nguyen
IC Ilsa M. Coleman
AK Arja Kaipainen
NS Navonil De Sarkar
PN Peter S. Nelson
CM Colm Morrissey
KK Keegan Korthauer
MP Mark M. Pomerantz
LE Leigh Ellis
BP Bogdan Pasaniuc
KL Kate Lawrenson
KK Kathleen Kelly
AZ Amina Zoubeidi
WH William C. Hahn
HB Himisha Beltran
HL Henry W. Long
MB Myles Brown
EC Eva Corey
MF Matthew L. Freedman
request Request a Protocol
ask Ask a question
Favorite

WCM154 organoids were cultured and maintained as described18. Organoids were dissociated to single cells using TrypLE (ThermoFisher). One million cells were resuspended in 20 μl of electroporation buffer (BTXpress) and mixed with 60 pmole of control or FOXA1 On-target pool siRNA (Dharmacon). Then organoid-siRNA mixtures were transferred to a 16-well NucleocuvetteTM Strip and nucleofection was performed in a 4D-Nucleofector (Lonza). Following nucleofection, 105 organoids cells were grown in a 12-well plate coated with 1% collagen I (ThermoFisher) for 7 days. Both adherent and floating cells were collected and stained with 0.4% trypan blue solution (ThermoFisher). Total cell numbers were measured by a hemocytometer. Cell proliferation with FOXA1 knock-down was normalized to control siRNA cells.

Copyright and License information: The Author(s) ©2021
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

post Post a Question
0 Q&A