The largest database of trusted experimental protocols

2 protocols using genticin

1

Generation of Knockout Mice via ES Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
Mitomycin C, Genticin, Penicillin, Streptomycine, Neomycin, M16 and M2 media, Plasmid purification kit, Tyrodes acid and leukemia Inhibitory Factor were purchased from Sigma (Germany). PMSG (Folligon) and hCG (Chorulon) were purchased from MSD Animal Health (New Zealand). DMEM, FCS and trypsin was obtained from Gibco BRL (France). RNA and DNA extraction kits were bought from Intron (Korea). Endofree Plasmid Maxi Kit was purchased from Qiagene (USA). High Pure PCR Purification Kit was from Roche (Germany). EW and R1 embryonic stem cells and pEGFP-C1 were kindly donated by Dr. Karim Nayernia at Institute of Molecular Medicine and Cell Therapy, Düsseldorf, Germany and GENEOCELL, Canada.
To generate knockout mouse, ES cells were cultured and electroporated with the gene targeting vector. The genetically-modified cells were selected and transferred into embryos. The modified ES cells along with the cells of embryo were used to produce chimeric mice. Crosses between the chimeras and wild type mice were resulted heterozygous offspring.
+ Open protocol
+ Expand
2

Overexpressing MafA in ADMSCs

Check if the same lab product or an alternative is used in the 5 most similar protocols
Differentiated ADMSCs were trypsinized and seeded
in 25 ml flasks 24 hours before the transfection. At day
10 of differentiation, cells were washed three times with
phosphate buffered saline (PBS, Calbiochem, Iran),
trypsinized, counted, and suspended at a density of 106/ml
in serum-free DMEM-HG. Then, 100 µl of cells were mixed
with 5 µg of suitable vector pCDNA(3.1+) in control group
and recombinant MafA/pCDNA 3.1(+) in the experimental
group in 0.4 ml electroporation cuvette (Biorad, USA), and
gently mixed by pipetting. The mixture was placed in an
electroporation system (GenePulser system II, Biorad, USA)
and one pulse of 140V was delivered for 15 milliseconds.
Following the electroporation, cells were plated onto a 25
ml flask that contained a differentiation medium and was
incubated at 37°C and 5% CO2. After 24 hours, Genticin
(350 µg/ml, Sigma, USA) was added to the growth media for
the positive selection of antibiotic-resistant ADMSCs. The
media were changed every 3 days, and Genticin selection
was maintained for 7 days. Antibiotic-resistant ADMSCs
were split to be grown for the differentiation.
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!