Stable Transfection/G418 (Geneticin) Selection Protocols
Summary: G418 (Geneticin) Selection. G418 is an aminoglycoside antibiotic produced by Micromonospora rhodorangea. G418 blocks polypeptide synthesis by inhibiting the elongation step in both prokaryotic and eukaryotic cells. Resistance to G418 is conferred by the neo gene from Tn5 encoding an aminoglycoside 3'-phosphotransferase, APT 3' II.
G418 vs Neomycin? - Cell Biology - Protocol Online
1998年10月28日 · G418 also known as geneticin, is a broad-spectrum antibiotic that will select mammalian cells expressing the neomycin protein (encoded by the neomycin gene). Neomycin will select bacteria expressing the same protein, but generally the promoter for Neo in mammalian expression plasmids is not usable in bacteria.
making G418 - Cell Biology - Protocol Online
2009年4月9日 · When G418 is produced it is not 100% active. The manufacturers determine the active fraction by bioassay. Your powder contains 736/1000 x 100 = 73.6% active G418. If you want to make a 50 mg/ml stock solution of ACTIVE G418 you therefore need to weigh out 1000/736 x 50= 67.93 mg of G418 for each ml of stock solution.
G418 Killing curve - Transfection and Transduction - Protocol Online
2007年3月15日 · Hi all, I have a little truble. I'm tring to select stable trasfected cell line (THP-1) with G418 . In order to determine the right concentration of G418 I'm doing a killing curve with non trasfected cells using a G418 concentration from 0.4 to 1mg .
G418 storage and contamination - Molecular Biology - Protocol …
I haven't had problems with storing G418 media for a few weeks at a time at 4 degrees. As long as the G418 and original media solutions are sterile to begin with, you shouldn't have problems. Contamination points to a handling problem at some stage in the preparation or use of the media, rather than a problem caused by the G418 itself.
HEK 293 Stables - Cell Biology - Protocol Online
2006年9月26日 · I have been working with HEK293 cells for a while and recently have been trying to generate stable cells expressing my protein of interest, unfortunately without success (using G418 selection). I have obtained transient expression of my protein of interest and have also been able to generate cells that stably express a fluorescent protein alone.
G418 Methods, Protocols and Troubleshootings
G418 and resistant cells - can't kill the cells (reply: 8) G418 selection protocol - need ideas for G418 selection (reply: 5) G418 effect on protein concentration - (reply: 1) false positive clones URGENT - resitant to G418 but not expressing protein (reply: 13) G418 Killing curve - How to determinate the right G418 concentration by killing curve ?
HEK293T and G418 - Cell Biology - Protocol Online
yes, HEK293T cells are G418 (Geneticin) resistant. They are stable transfectants with an aph3' resistance cassette. So you can not select for your plasmid. I would suggest that you replace your NeoR with a Hygromycin resistance cassette. Usual concentration we use is 200 ug/ml for selection and 100 ug/ml for maintaining the cells.
hek293t and geneticin G418 - Cell Biology - Protocol Online
hek293t and geneticin G418 - (Mar/01/2007 ) Hi, I know that this has been addressed to some extent before but I am still not clear so would love some advice!! I was wondering if anyone can tell me whether HEK293T cells are resistant to Geneticin (G418) in general.
false positive clones URGENT - Transfection and Transduction
2007年3月28日 · for g418 selection, just follow the protocol. you should use the lower amount g418 wichi can kill all of cells within 1-2 weeks. High amount g418 and killed the cells ASAP does not mean good.