Editing Bacterial Strains for Protein Expression (section)

== Bacterial Strains for Protein Expression  ==

{| border="1"
|-
| '''Bacterial Strain''' 
| '''Company''' 
| '''Features''' 
| '''Growth Requirements'''
|-
| BL21(DE3)&nbsp; 
| Novagen 
| DE3 lysogen contains T7 polymerase upon IPTG induction. This strain is defficient of lon and omp-t proteases and is therefore suitable for expression of non-toxic genes. 
| There is no tight control over the expression, thus addition of 1% glucose to the growth medium should be considered. (<u>Note</u>: '''1.''' This significantly attenuates growth rates. '''2.''' Higher concentrations of IPTG should be used for induction.)
|-
| BL21(DE3)-pLysS&nbsp; 
| Novagen 
| DE3 lysogen expresses T7 polymerase upon IPTG induction.&nbsp; The pLysS plasmid produces T7 lysozyme to reduce basal level expression of the gene of interest. Thus it is suitable for&nbsp; expression of toxic genes.&nbsp; [''pLysS contains the p15A origin. This origin allows pLysS to be compatible with plasmids containing the ColE1 or pMB1 origin (i.e. pUC- or pBR322- derived plasmids)''.] 
| Chloramphenicol 34ug/ml&nbsp;
|-
| BL21 Star-pLysS&nbsp; 
| Invitrogen 
| This strain is mutated in Rnase E,&nbsp; thus Rnase degradation is reduced, and protein expression may be increased. The pLysS presence allows tight control of expression. Use this strain for both soluble and insoluble protein production. 
| Chloramphenicol 34ug/ml&nbsp;
|-
| BL21-SI&nbsp; 
| Invitrogen 
| T7 polymerase in the DE3 is under control of the salt-inducible proU&nbsp; promoter. Induction of protein production at 0.1-0.5M of NaCl. Helps protein solubility. 
| '''LBON''' medium (LB w/o NaCl), '''30<sup>o</sup>c'''.&nbsp; Expression induction by NaCl, if controlled by a T7 promoter. Otherwise - IPTG.
|-
| BL21-AI&nbsp; 
| Invitrogen 
| BL21-AI was constructed by inserting a chromosomal copy of the T7 RNA polymerase gene under the tight control of the arabinose-inducible araBAD promoter.&nbsp; 
| Tetracyclin 12.5ug/ml&nbsp; Expression induction by arabinose. (If using pET vectors, IPTG is also required!) Glucose will repress this induction.
|-
| Tuner&nbsp; 
| Novagen 
| Contains a mutation in the lac permease (lacZY) gene. This enables adjustable levels of protein expression throughout all cells in a culture. The lac permease (lacY) mutation allows uniform entry of IPTG into all cells in the population, which produces a concentration-dependent, homogeneous level of induction. By adjusting the concentration of IPTG, expression can be regulated from very low levels up to the robust, fully induced levels commonly associated with pET vectors. Lower level expression may enhance the solubility and activity of difficult target proteins. 
| None&nbsp;
|-
| Tuner pLysS&nbsp; 
| Novagen 
| Contains the pLysS plasmid (tighter control over expression) in addition to the lac permease mutation. 
| Chloramphenicol 34ug/ml&nbsp;
|-
| Origami&nbsp; 
| Novagen 
| Origami host strains are K-12 derivatives that have mutations in both the thioredoxin reductase (trxB) and glutathione reductase (gor) genes, which greatly enhances disulfide bond formation in the cytoplasm.&nbsp; 
| Kanamycin 15ug/ml&nbsp; Tetracyclin 12.5ug/ml&nbsp;
|-
| Origami B&nbsp; 
| Novagen 
| Origami B host strains carry the same trxB/gor mutations as the original Origami strains, except that they are derived from a lacZY mutant of BL21. Thus the Origami B strains combine the desirable characteristics of BL21, Tuner&amp;trade; and Origami hosts in one strain background. 
| &nbsp;Kanamycin 15ug/ml&nbsp; Tetracyclin 12.5ug/ml&nbsp;
|-
| Origami B pLysS&nbsp; 
| Novagen 
| &nbsp; 
| 
Kanamycin 15ug/ml&nbsp; 

Tetracyclin 12.5ug/ml 

Chloramphenicol 34ug/ml&nbsp; 

|-
| Rosetta&nbsp; 
| Novagen 
| Rosetta host strains are BL21 lacZY (Tuner&amp;trade;) derivatives designed to enhance the expression of eukaryotic proteins that contain codons rarely used in E. coli. These strains supply tRNAs for the codons AUA, AGG, AGA, CUA, CCC, GGA on a compatible chloramphenicol resistant plasmid. The tRNA genes are driven by their native promoters. 
| Chloramphenicol 34ug/ml&nbsp;
|-
| Rosetta pLysS&nbsp; 
| Novagen 
| &nbsp;In Rosetta(DE3)pLysS the rare tRNA genes are present on the same plasmids that carry the T7 lysozyme. 
| Chloramphenicol 34ug/ml&nbsp;
|-
| Rosetta-gami-pLysS&nbsp; 
| Novagen 
| Rosetta-gami host strains are Origami derivatives that combine the enhanced disulfide bond formation resulting from trxB/gor mutations with enhanced expression of eukaryotic proteins&nbsp; that contain codons rarely used in E. coli. These strains supply tRNAs for AGG, AGA, AUA, CUA, CCC, GGA on a compatible chloramphenicolresistant plasmid. The tRNA genes are driven by their native promoters. In Rosetta gami(DE3)pLysS the rare tRNA genes are present on the same plasmids that carry the T7 lysozyme.&nbsp; 
| 
Kanamycin 15ug/ml 

Tetracyclin 12.5ug/ml&nbsp; 

Chloramphenicol 34ug/ml&nbsp; 

|-
| BL21 CodonPlus&nbsp; 
| Stratagene 
| BL21-CodonPlus-'''RIL''' chemically competent cells carry extra copies of the argU, ileY, and leuW tRNA genes. The tRNAs encoded by these genes recognize the '''AGA/AGG''' ('''arginine'''), '''AUA''' ('''isoleucine'''), and '''CUA''' ('''leucine''') codons, respectively.&nbsp; 
| Tetracyclin 12.5ug/ml Chloramphenicol 34ug/ml
|-
| AD494&nbsp; 
| Novagen 
| AD494 strains are thioredoxin reductase (trxB) mutants of the K12 strain that enable disulfide bond formation in the cytoplasm, providing the potential to produce properly folded active proteins. 
| &nbsp;Kanamycin 15ug/ml&nbsp;
|-
| BL21trxB&nbsp; 
| Novagen 
| &nbsp;BL21trxB strains possess the same thioredoxin reductase mutation (trxB) as the AD494 strains in the protease deficient BL21 background. The trxB mutation enables cytoplasmic disulfide bond formation. 
| &nbsp;Kanamycin 15ug/ml&nbsp;
|-
| HMS174&nbsp; 
| Novagen 
| HMS174 strains provide the recA mutation in a K-12 background. Like BLR, these strains may stabilize certain target genes whose products may cause the loss of the DE3 prophage. 
| None&nbsp;
|-
| NovaBlue(DE3)&nbsp; 
| Novagen 
| NovaBlue(DE3) is a K-12 strain ideally suited as an initial cloning and expression host due to its high transformation efficiency&nbsp; and recA endA mutations, which result in high yields of excellent quality plasmid DNA. It olso contains DE3 for T7 polymerase expression. Ttight regulation of expression is conferred due to the presence of lacI<sup>q </sup>repressor encoded by the F episome. 
| Tetracyclin 12.5ug/ml&nbsp;
|-
| BLR&nbsp; 
| Novagen 
| BLR is a recA<sup>-</sup> derivative of BL21 that improves plasmid monomer yields and may help stabilize target plasmids containing repetitive sequences or whose products may cause the loss of the DE3 prophage. 
| Tetracyclin 12.5ug/ml&nbsp;
|-
| C41(DE3) 
| Lucigen 
| Effective in expressing toxic and membrane proteins from all classes of organisms,including viruses,eubacteria,archaea,yeasts,plants,insects,and mammals. The strain C41(DE3)was derived from BL21(DE3) [E.coli F ompT hsdS B (r B -m B -) gal dcm (DE3)].This strain has at least one uncharacterized mutation that prevents cell death associated with expression of many toxic recombinant proteins. 
| None
|-
| C43(DE3) 
| Lucigen 
| Effective in expressing toxic and membrane proteins from all classes of organisms,including viruses,eubacteria,archaea,yeasts,plants,insects,and mammals. 
| None
|}

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'''Rosetta 2''' host strains are BL21 derivatives designed to enhance the expression of eukaryotic proteins that contain codons rarely used in ''E. coli''. These strains supply tRNAs for 7 rare codones (AGA, AGG, AUA, CUA, GGA, CCC, and CGG) on a compatible chloramphenicol-resistant plasmid. The tRNA genes are driven by their native promoters. 

'''DE3''' indicates that the host is a lysogen of DE3, and therefore carries a chromosomal copy of the T7 RNA polymerase gene under control of the ''lacUV5'' promoter. Such strains are suitable for production of protein from target genes cloned in pET vectors by induction with IPTG. 

'''pLysS''' strains express T7 lysozyme, which further suppresses basal expression of T7 RNA polymerase prior to induction, thus stabilizing pET recombinants encoding target proteins that affect cell growth and viability. '''Genotype: '''F<sup>-</sup> ''ompT hsdS''<sub>B</sub>(r<sub>B</sub><sup>-</sup> m<sub>B</sub><sup>-</sup>) ''gal dcm'' (DE3) pLysSRARE2 (Cam<sup>R</sup>)<br> 

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