How do you transform cells in BL21?

BL21 Chemically Competent Cells are transformed in 40 μL reactions. Cells are packaged with sufficient volumes for 1, 2, or 4 reactions per tube. Transformation is performed by heat shock at 42 °C, followed by incubation on ice.

Can you isolate plasmid from BL21?

BL21 cells are not designed for plasmid purification but protein expression. BL21 lacks in some proteases but has the endonucleases present. You can make mini-preps and try to isolate the plasmid, but there is a risk that your plasmid may be digested.

What are BL21 cells?

BL21 competent cells are an all-purpose strain for high-level protein expression and easy induction. The basic BL21 strain does not contain the T7 RNA polymerase gene and can be used with non-T7 RNA polymerase protein expression systems. BL21 has the tightest control of protein expression for extremely toxic proteins.

Why is DH5alpha used?

DH5 alpha has a recA mutation, so it does no heterologous recombination which ensures a higher insert stability . Additionally, it lacks some endonucleases which might digest the plasmids during the isoation procedure. DH5 alpha is additionally competent for blue-white screening.

What is a BL21 cell?

Thermo Scientific™BL21(DE3) Competent Cells are suitable for the expression of nontoxic heterologous genes. The strain contains the DE3 lysogen that carries the gene for T7 RNA polymerase under control of a lacUV5 promoter, allowing expression of the T7 RNA polymerase to be induced with IPTG.

How do you prepare DNA from E coli BL21?

(For C2527I) Thaw a tube of BL21 (DE3) Competent E. coli cells on ice until the last ice crystals disappear. Mix gently and carefully pipette 50 µl of cells into a transformation tube on ice. Add 1–5 µl containing 1 pg–100 ng of plasmid DNA to the cell mixture. Carefully flick the tube 4–5 times to mix cells and DNA.

What is BL21 DE3 competent?

The BL21(DE3) competent cells are an all-purpose strain for high-level protein expression and easy induction. The BL21(DE3)pLysS competent cells provide tighter control of protein expression for expression of toxic proteins and are resistant to chloramphenicol.

What is the genotype of bl21-ai?

Genotype BL21-AI™: F- ompT hsdSB (rB-mB-) gal dcm araB::T7RNAP- tetA The BL21-AI™ strain is an E. coli B/r strain and does not contain the lon protease. It is also deficient in the outer membrane protease, OmpT. The lack of these proteases reduces degradation of heterologous proteins expressed in this strain.