A549 Cells GFP-SMAD4,

Application

To learn more, please visit the Cellular Reporter Cell Line webpage

This product is a human A549 cell line in which the genomic SMAD4 gene has been endogenously tagged with a Green Fluorescent Protein (GFP) gene using CompoZr^® zinc finger nuclease (ZFN) technology. Integration resulted in endogenous expression of the fusion protein in which GFP is attached to the C-terminus of SMAD4. Fluorescence imaging shows SMAD4-GFP expression throughout the cell. Upon activation with a ligand such as transforming growth factor beta (TGFb), the cell line shows redistribution of SMAD4 from the cytoplasm to the nucleus, making it useful for high content screening to identify compounds that modulate this redistribution. This stable cell line was expanded from a single clone. The target’;s gene regulation and corresponding protein function are preserved in contrast to cell lines with overexpression via an exogenous promoter.

Features and Benefits

Zinc Finger Nuclease (ZFN)-mediated targeted integration of a fluorescent GFP tag into the last coding exon just preceding the stop codon of the SMAD4 gene on chromosome 18q21.2 to create a cell line exhibiting stable expression of the GFP transgene.The A549 cells are adherent, with a doubling time of approx. 22 hours.

General description

A549 GFP-SMAD4 are lung carcinoma cells from a human 58 year old caucasian male having two distinct ZFN modifications creating two fluorescent transgenes each expressed from their endogenous gene locus.This cell line was derived from ATCC Catalog No. CCL-185.

Legal Information

CompoZr is a registered trademark of Sigma-Aldrich Co. LLC

Preparation Note

Media Renewal changes two to three times per week.Rapidly thaw vial by gentle agitation in 37°C water bath (~2 minutes), keeping vial cap out of the water. Decontaminate with 70% ethanol, add 9 mL culture media and centrifuge 125 x g (5-7 minutes). Resuspend in complete culture media and incubate at 37°C in a 5% CO2 atmosphere.Subculture Ratio: approx. 1:3-1:8The base medium for this cell line is F12 Ham Medium, Cat. No. N4888. To make the complete growth medium, add the following components to the base medium: fetal bovine serum, Cat. No. F4135, to a final concentration (v/v) of 10% and L-glutamine, Cat. No. G7513, at a final concentration of 2 mM.Cell freezing medium-DMSO 1X, Cat. No. C6164.

Quality

Tested for Mycoplasma, sterility, post-freeze viability, short terminal repeat (STR) analysis for cell line identification, cytochrome oxidase I (COI) analysis for cell line species confirmation.