RNA Extraction Assay: A Method to Extract RNA from miRNA Transfected Lung Cancer Cells

Published: April 30, 2023

Abstract

Source: Hossian, A. K. M. N. et al., Analysis of Combinatorial miRNA Treatments to Regulate Cell Cycle and Angiogenesis. J. Vis. Exp. (2019).

This video describes the RNA extraction protocol from miRNA transfected lung cancer cells. This approach can be used to identify a combinatorial miRNA treatment activity in halting the cell cycle and angiogenesis.

Protocol

1. miR-143 and miR-506 transfection CAUTION: Use latex gloves, protective eyeglasses, and a laboratory coat while performing the described experiments. When required, use the biosafety cabinet with the cabinet fan on, without blocking the airways or disturbing the laminar airflow. Always set the protecting glass window to the appropriate height, as described by the manufacturer. Seed NSCLC A549 cells in a T25 cm2 flask/6/96 well plate in…

開示

The authors have nothing to disclose.

Materials

A549 Non Small Cell Lung Cancer Cells ATCC ATCC CCL-185
Ambion Silencer Negative Control No. 1 siRNA Ambion AM4611
hsa-miR-143-3p miRNA Mimic ABM MCH01315
hsa-miR-506-3p miRNA Mimic ABM MCH02824
Fetal Bovine Serum (FBS) Fisher Scientific 10438026
HyClone Phosphate Buffered Saline (PBS) Fisher Scientific SH30256FS
DMEM 4.5 g/L Glucose, w/out Sodium Pyruvate, w/ L-Glutamine VWR VWRL0100-0500
Invitrogen UltraPure DNase/ RNase-Free Distilled Water Invitrogen 10-977-015
Quick-RNA Kits Zymo Research R1055
Ethanol, Absolute (200 Proof), Molecular Biology Grade, Fisher BioReagents BP2818500
-80 °C Freezer VWR VWR40086A
Nanodrop Thermo Scientific NanoDrop one C
96 well plate CELLTREAT Scientific 50-607-511
Penicillin-streptomycin 10/10 Antlanta Biologicals B21210
CO2 Incubator Thermo Scientific HERAcell 150i
dnase Zymo Research E1010
F-12K Nutrient Mixture (Kaighn's Mod.) with L-glutamine, Corning Corning 45000-354
Fetal Bovine Serum – Premium Antlanta Biologicals S11150
Temperature controlled centrifuge matchine Thermo Scientific ST16R
Temperature controlled micro centrifuge matchine Eppendrof 5415R

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記事を引用
RNA Extraction Assay: A Method to Extract RNA from miRNA Transfected Lung Cancer Cells. J. Vis. Exp. (Pending Publication), e20237, doi: (2023).

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