| Criteria | BL21 | BL21(DE3) | BL21(DE3)pLysS | Remarks |
| Genetic Background | Wild-type E. coli B strain derivative | BL21 containing the λDE3 lysogen, which harbors the T7 RNA polymerase gene | BL21(DE3) carrying the pLysS plasmid, encoding T7 lysozyme | All are derivatives of BL21, differing in ability to express proteins via T7 system. |
| T7 RNA Polymerase | ❌ Not present | ✅ Present in the genome, under control of lacUV5 promoter | ✅ Present (same as BL21(DE3)) | T7 polymerase enables high-level expression from T7 promoter-based vectors. |
| Expression Vector Compatibility | Incompatible with T7-based vectors without helper plasmid | ✅ Compatible with T7-promoter vectors (e.g., pET series) | ✅ Compatible with T7-promoter vectors | BL21 alone requires other systems (e.g., lac, trc) for expression. |
| Basal Expression Level | No expression from T7 promoter | Moderate basal expression possible (leaky) | Very low basal expression due to T7 lysozyme inhibition | pLysS strain is ideal for expressing toxic or unstable proteins. |
| pLysS Plasmid | ❌ Absent | ❌ Absent | ✅ Present – encodes T7 lysozyme (inhibits T7 RNA polymerase) | Requires chloramphenicol selection due to presence of the pLysS plasmid. |
| Inducer for Expression | Depends on vector used (e.g., lac, tac promoters) | IPTG (induces T7 RNA polymerase) | IPTG (same mechanism) | All DE3 derivatives use IPTG to induce T7 expression. |
| Protein Yield | Low to moderate (non-T7 based) | High with T7 vectors | Moderate to high; slightly reduced due to T7 inhibition, but more controlled | Slightly reduced yield in pLysS, but with improved control over expression. |
| Suitability for Toxic Protein Expression | Not suitable | Less suitable – leaky expression may harm cells | Most suitable – tight control reduces toxicity risk | pLysS offers best safety margin for unstable or toxic recombinant proteins. |
| Antibiotic Resistance | None (unless from plasmid) | None (unless from plasmid) | Chloramphenicol resistance (from pLysS plasmid) | Consider antibiotic selection pressure in co-transformations. |
| Applications | General cloning, basic protein expression | High-level expression of recombinant proteins with T7-based systems | Expression of toxic or tightly regulated proteins using T7-based vectors | Selection depends on expression strength vs. control need. |
