E. coli Expression Strains

NEB offers several stains with varying levels of expression control, each with phage T1 resistance and extremely high transformation efficiencies.

NEB Express is an enhanced BL21 derivative available with (NEB #C3037) or without (NEB #C2523) the added control from lacIq.

Within the realm of E. coli expression, the T7 system is the most popular approach for producing proteins. In this system, an expression vector containing a gene of interest cloned downstream of the T7 promoter is introduced into a T7 expression host. T7 expression hosts, such as DE3 strains or T7 Express strains, carry a chromosomal copy of the phage T7 RNA Polymerase gene, which is controlled by a lac promoter. When inducer is added, T7 RNA Polymerase is expressed and becomes dedicated to transcription of the gene of interest. T7 Express strains are available with the control features lacIq and/or lysY. The lysY gene expresses a variant T7 lysosome which naturally inhibits T7 RNA Polymerase, thereby controlling basal expression of the target gene. LysY is expressed from a single copy mini-F plasmid that does not require antibiotic selection for propagation. T7 Express lysY/Iq (NEB #C3013) provides the highest level of expression control.

BL21(DE3) (NEB #C2527) is available for routine T7 expression and Lemo21(DE3) (NEB #2528) is designated for tunable T7 expression of difficult targets such as membrane proteins, toxic proteins and proteins prone to insoluble expression. NiCo21(DE3) (NEB #C2529) has been recently engineered to improve the recovery of His-tagged proteins.

SHuffle® strains enable disulfide bond formation in the cytoplasm. Normally, reductases in the E. coli cytoplasm keep cysteines in their reduced form, thereby preventing any disulfide bond formation in this compartment. The deletions of the genes for glutaredoxin reductase and thioredoxin reductase (Δgor ΔtrxB), allows disulfide bond formation in the cytoplasm with SHuffle. This combination of mutations is normally lethal, but the lethality is suppressed by a mutation in the peroxiredoxin enzyme (ahpC*). In addition, SHuffle expresses a version of the periplasmic disulfide bond isomerase, DsbC, which lacks its signal sequence, retaining it in the cytoplasm. This enzyme has been shown to act on proteins with multiple disulfide bonds, to correct mis-oxidized bonds and promote proper folding . The gene for the cytoplasmic DsbC is present on the chromosome.

Available E. coli Expression Strains:

Strain	NEB #	Characteristics
BL21 Competent E. coli	C2530	Routine non-T7 expression
BL21(DE3) Competent E. coli	C2527	Routine T7 expression
NEB Express Competent E. coli	C2523	Versatile non-T7 expression strain Protease deficient
NEB Express Iq Competent E. coli	C3037	Control of IPTG induced expression from Plac, Ptac and Ptrc Protease deficient
T7 Express Competent E. coli	C2566	Most popular T7 expression strain Protease deficient
T7 Express Iq Competent E. coli	C3016	T7 expression Protease deficient Reduced basal expression
T7 Express lysY Competent E. coli	C3010	T7 expression Protease deficient Better reduction of basal expression
T7 Express lysY/Iq Competent E. coli	C3013	T7 expression Protease deficient Highest level of expression control
T7 Express Crystal Competent E. coli	C3022	T7 expression Protease deficient SeMet labeling for protein crystallography
SHuffle Express Competent E. coli	C3028	Protease deficient/B strain Enhanced capacity to correctly fold proteins with multiple disulfide bonds in the cytoplasm
SHuffle T7 Express Competent E. coli	C3029	T7 expression Protease deficient/B strain Enhanced capacity to correctly fold proteins with multiple disulfide bonds in the cytoplasm
SHuffle T7 Express lysY Competent E. coli	C3030	T7 expression Protease deficient/B strain Tightly controlled expression of toxic proteins Enhanced capacity to correctly fold proteins with multiple disulfide bonds in the cytoplasm
SHuffle T7 Competent E. coli	C3026	T7 expression/K12 strain Enhanced capacity to correctly fold proteins with multiple disulfide bonds in the cytoplasm
NiCo21(DE3) Competent E. coli	C2529	Improved purity of target proteins isolated by IMAC
Lemo21(DE3) Competent E. coli	C2528	Tunable T7 expression for difficult targets