What is the difference between the selectable marker and the antibiotic resistance gene in the Addgene plasmid?

Usually when Addgene says “bacterial resistance” it means “selection for the plasmid in E. coli for cloning/plasmid maintenance” and “selectable marker” means it’s an active marker when transferred to the organism of interest.

How do you transfect a plasmid into a mammalian cell?

Plasmid transfection into mammalian cells is fairly straightforward and the resultant cells can either express the plasmid DNA transiently (similar to bacteria) or incorporate the genetic material directly into the genome to form a stable transfection.

Can plasmids replicate in mammalian cells?

Few naturally-occurring plasmids are maintained in mammalian cells. These two genomes are replicated in a licensed manner, each using a single viral protein and cellular replication machinery, and are passed to daughter cells during cell division despite their lacking traditional centromeres (4-8).

Which are the most commonly used selectable markers?

Commonly used selectable marker systems include neomycin phosphotransferase, hygromycin phosphotransferase, phosphoinothricin acetyltransferase, and glyphosate oxidoreductase. Use of nptII in concert with the antibiotic kanamycin has become the most widely used selectable marker gene system in dicotyledonous plants.

Which of the following antibiotic resistance genes are used as selectable marker for E coli?

Normally the genes encoding resistance to antibiotics such as ampicillin, chloroamphenicol, tetracycline or kanamycin, etc., are considered useful selectable markers for E. coli.

Why are antibiotic resistance genes used as markers in E coli?

Adding an antibiotic resistance gene to the plasmid solves both problems at once – it allows a scientist to easily detect plasmid-containing bacteria when the cells are grown on selective media, and provides those bacteria with a pressure to keep your plasmid. Viva la (bacterial) resistance!

Which of the following is the most efficient method for transfecting mammalian cells?

electroporation
Basically, electroporation is the most efficient method for cell transfection.

Does plasmid DNA integrate into genome?

1. In a stable transfection, does the plasmid DNA always get integrated into the genome? Unlike plasmids used for expression in yeast and bacteria, plasmids used for stable transfections rarely contain an ORI since the integrated DNA will be replicated as part of the genome.

Can you transform mammalian cells?

Mammalian cells will take up and express genes when they are exposed to either metaphase chromosomes or naked genomic or recombinant DNA. In many cases the transforming DNA is stably expressed, and consequently alters the genotype of the recipient cell.

Why a cloning vector must have a selectable marker?

Selectable markers are essential to identify and eliminate non-transformants(no recombinant DNA), and selectively permitting the growth of the transformants (host cells bearing recombinant DNA).

How are selectable markers used in cell selection?

A subset of each of these can be amplified, or the copy number of the marker gene can be further increased, by application of the proper selective agent (50,51). Many of the dominant selectable markers are bacterial antibiotic resistant genes or genes that detoxify the effects of various protein synthesis inhibitors on the cell.

How to select a selectable mammalian expression vector?

Selectable mammalian expression vectors. The purified plasmids were transfected into HEK239 cells by lipofection (Lipofectamine 2000) or nucleofection (Lonza), with a >90% transfection ratio in both cases. Survival rates of cells transfected with the eluted DNA were compared to controls containing no added DNA.

Which is antibiotic resistance marker is expressed separately from SV40 promoter?

The antibiotic resistance marker, either puromycin or neomycin, is expressed separately from an SV40 promoter. pBApo-CMV lacks a selectable marker. Cat. # pBApo-CMV is a plasmid for gene expression in mammalian cells.

Which is a selectable vector for pbapo-CMV?

The pBApo series of vectors are mammalian expression vectors for constitutive expression of your gene of interest from either a CMV or EF1-alpha promoter. The antibiotic resistance marker, either puromycin or neomycin, is expressed separately from an SV40 promoter. pBApo-CMV lacks a selectable marker.