{"id":86,"date":"2017-04-19T15:52:36","date_gmt":"2017-04-19T20:52:36","guid":{"rendered":"https:\/\/www.vanderbilt.edu\/csb\/?page_id=86"},"modified":"2024-08-28T15:01:57","modified_gmt":"2024-08-28T20:01:57","slug":"e-coli-strains","status":"publish","type":"page","link":"https:\/\/www.vanderbilt.edu\/csb\/facilities\/labs-instrumentation-facility\/protein-characterization\/e-coli-strains\/","title":{"rendered":"E. coli Strains"},"content":{"rendered":"

Host strains<\/h2>\n

Cells are available to VU researchers. If you are interested in any of the E.coli<\/em> strains listed below, please contact arina.hadzi@vanderbilt.edu<\/a>.<\/p>\n

\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n
Strain<\/th>\nSource<\/th>\nResistance<\/th>\nComments<\/th>\n<\/tr>\n
<\/td>\n<\/td>\n<\/td>\n<\/td>\n<\/tr>\n
ArcticExpress(DE3)<\/td>\nAgilent Technologies (formerly Stratagene)<\/td>\nNone*<\/td>\nContains genes for cold-adapted chaperonins (cpn60, cpn10) to enhance folding\/growth at low temperatures and increase soluble protein yields. See the expression protocol instruction manual<\/a>. Co-purification of cpn60\/10 with the protein of interest is a common problem. Incubation with MgCl2\/ATP\/KCl facilitates separation. See: R. E. Joseph; A. H. Andreotti, Protein Expr. Purif. 2008, 60, 194\u2013197<\/a>.<\/td>\n<\/tr>\n
ArcticExpress(DE3)RIL<\/td>\nAgilent Technologies (formerly Stratagene)<\/td>\nNone**<\/td>\nContains genes for cold-adapted chaperones and tRNAs for rare Arg\/Ile\/Leu codons. See the expression protocol instruction manual<\/a>. See above note on purification strategy.<\/td>\n<\/tr>\n
BL21(DE3)<\/td>\nAgilent Technologies (formerly Stratagene)<\/td>\nNone<\/td>\nGeneral purpose T7 expression strain.<\/td>\n<\/tr>\n
BL21(DE3)pLysS<\/td>\nAgilent Technologies (formerly Stratagene)<\/td>\nCap<\/td>\nContains gene for T7 lysozyme. T7 lysozyme inhibits T7 RNA polymerase and allows tighter expression control.<\/td>\n<\/tr>\n
BL21(DE3)Star<\/td>\nLife Technologies (formerly Invitrogen)<\/td>\nNone<\/td>\nContains mutation in RNaseE to improve stability of mRNA transcripts and increase yield.<\/td>\n<\/tr>\n
BL21CodonPlus(DE3)RIL<\/td>\nAgilent Technologies (formerly Stratagene)<\/td>\nCap<\/td>\nContains tRNAs for rare Arg (AGA, AGG), Ile (AUA), and Leu (CUA) codons that often restrict translation from AT-rich genomes.<\/td>\n<\/tr>\n
BL21CodonPlus(DE3)RP<\/td>\nAgilent Technologies (formerly Stratagene)<\/td>\nCap<\/td>\nContains tRNAs for rare Arg (AGA, AGG) and Pro (CCC) codons that often restrict translation from GC-rich genomes.<\/td>\n<\/tr>\n
C41(DE3)<\/td>\nSanders lab<\/td>\nNone<\/td>\nMutant BL21 strain shown to be effective for expression of toxic and membrane proteins.<\/td>\n<\/tr>\n
C41(DE3)pLysS<\/td>\nMade in-house from C41(DE3)<\/td>\nCap<\/td>\nMutant BL21 strain shown to be effective for expression of toxic and membrane proteins. Contains pLysS for tighter expression control.<\/td>\n<\/tr>\n
C43(DE3)<\/td>\nSanders lab<\/td>\nNone<\/td>\nMutant BL21 strain shown to be effective for expression of toxic and membrane proteins.<\/td>\n<\/tr>\n
Origami B(DE3)<\/td>\nEMD Millipore (formerly Novagen)<\/td>\nKan\/Tet<\/td>\nBL21 derivative with mutant thioredoxin reductase (trxB) and glutathione reductase (gor) genes to enhance cytosolic disulfide bond formation. Use of a thioredoxin fusion tag further enhances the formation of disulfide bonds in the cytoplasm.<\/td>\n<\/tr>\n
Rosetta2(DE3)<\/td>\nEMD Millipore (formerly Novagen)<\/td>\nCap<\/td>\nEnhances the expression of eukaryotic proteins that contain codons rarely used in E. coli<\/i>. Contains tRNAs for 7 rare codons (AUA, AGG, AGA, CUA, CCC, GGA, CGG).<\/td>\n<\/tr>\n
Rosetta2(DE3)pLysS<\/td>\nEMD Millipore (formerly Novagen)<\/td>\nCap<\/td>\nEnhances the expression of eukaryotic proteins that contain codons rarely used in E. coli<\/i>. Contains tRNAs for 7 rare codons and pLysS for tighter expression control.<\/td>\n<\/tr>\n
SoluBL21(DE3)<\/a><\/td>\nGenlantis<\/td>\nNone<\/td>\nImproves expression of toxic clones and proteins in soluble form.
\nOnly available from a glycerol stock.<\/td>\n<\/tr>\n
Tuner(DE3)<\/td>\nEMD Millipore (formerly Novagen)<\/td>\nNone<\/td>\nBL21 lacZY deletion that enables tuning expression levels by adjusting IPTG concentration.<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

Cap=Chloramphenicol; Kan=Kanamycin; Tet=Tetracycline
\n*Chaperonin-encoding genes are on a gentamycin (Gent) plasmid. You don’t need to add Gent to transformation plates, but vendor recommends selection when expressing protein.
\n**Chaperonin-encoding genes are on a Gent plasmid (see above). Rare tRNA genes are on a streptomycin (Strep) plasmid with a functional partitioning locus that prevents plasmid loss even in the absence of Strep.
\n***Rare Arg, Ile, Leu tRNA genes are on a streptomycin (Strep) plasmid with a functional partitioning locus that prevents plasmid loss even in the absence of Strep.<\/span><\/p>\n

Transformation Protocol:<\/b>
\nUse for all strains except<\/u> SoluBL21(DE3). For SoluBL21 protocol, click on the link in table above.
\nCell aliquots are 100 uL. Can use as little as 20 uL per transformation.
\nNOTE: The 45 sec heat shock was found to be optimal. Using LB in lieu of SOC often results in fewer colonies.<\/span><\/p>\n

    \n
  • Thaw cells on ice.<\/span><\/li>\n
  • Add 10 ng DNA per 50 uL of cells.<\/span><\/li>\n
  • Flick the tube several times to ensure even distribution of DNA.<\/span><\/li>\n
  • Ice for 20 min.<\/span><\/li>\n
  • Heat shock cells for 45 sec at 42 \u00b0C.<\/span><\/li>\n
  • Ice for 2 min.<\/span><\/li>\n
  • Add 450 uL of SOC medium and incubate for 1 hour at 37 \u00b0C with shaking.<\/span><\/li>\n
  • Plate 100 uL of transformation mix on antibiotic plates.<\/span><\/li>\n<\/ul>\n

     <\/p>\n

     <\/p>\n","protected":false},"excerpt":{"rendered":"

    Host strains Cells are available to VU researchers. If you are interested in any of the E.coli strains listed below, please contact arina.hadzi@vanderbilt.edu. Strain Source Resistance Comments ArcticExpress(DE3) Agilent Technologies (formerly Stratagene) None* Contains genes for cold-adapted chaperonins (cpn60, cpn10) to enhance folding\/growth at low temperatures and increase soluble protein yields. See the expression protocol…<\/p>\n","protected":false},"author":285,"featured_media":0,"parent":80,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"page_vunetid.php","meta":{"spay_email":""},"tags":[],"acf":[],"jetpack_sharing_enabled":true,"_links":{"self":[{"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/pages\/86"}],"collection":[{"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/users\/285"}],"replies":[{"embeddable":true,"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/comments?post=86"}],"version-history":[{"count":18,"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/pages\/86\/revisions"}],"predecessor-version":[{"id":5556,"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/pages\/86\/revisions\/5556"}],"up":[{"embeddable":true,"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/pages\/80"}],"wp:attachment":[{"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/media?parent=86"}],"wp:term":[{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.vanderbilt.edu\/csb\/wp-json\/wp\/v2\/tags?post=86"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}