Plasmids 101: Common Lab E. coli Strains

By Matthew Ferenc


This post was updated on Nov 14, 2017.

You've worked hard designing your plasmid – you carefully selected the optimal promoter for your gene of interest, painstakingly cloned into the perfect empty backbone, made sure to add the right tags to your gene, and may have even put a fluorescent protein downstream, separated by an IRES element. You did a lot of work! But let’s take a moment to recognize your little prokaryotic minions that carried out the labor-intensive process of replicating your new plasmid: the Escherichia coli bacteria.

It’s hard to count the number of different commercial strains of E. coli currently available  a quick Google search suggests there are hundreds. This only includes general lab strains designed for subcloning or protein expression. If you were to include customized strains, the number is probably in the thousands! The goal of this article is to provide enough background for you to distinguish the features of any common lab strain and determine whether it is appropriate for propogating your plasmid or carrying out your experiment.

History of E. coli strains

E. coli are gram-negative, rod shaped bacteria that were named after Dr. Theodor Escherich, the scientist who first described them in 1885. E. coli are mainly found in the intestinal tract of animals. There are many different naturally occurring strains of E. coli, some of which are deadly to humans. The majority of all common, commercial lab strains of E. coli used today are descended from two individual isolates, the K-12 strain and the B strain. K-12 was isolated from a patient in 1920 and eventually led to the common lab strains MG1655 and its derivatives DH5alpha and DH10b (also known as TOP10). The history of B strain is a bit more convoluted due to researchers sharing and renaming samples throughout history. It was likely isolated in 1918 but was first referred to as “B strain” in 1942. The BL21 strain and derivatives are the most common examples of the E. coli B strain.

Common E. coli strains used in the lab

Bacterial plate

Most of the commercial strains you find today are marketed for a specific purpose: fast growth, high-throughput cloning, routine cloning, cloning unstable DNA, preparing unmethylated DNA, and more. Many mutations that make these features possible are present in most commercial strains, especially mutations that make major improvements such as those that increase plasmid yield and/or DNA quality. Table 1 below outlines a few of the more common genetic changes found in E. coli strains.

Table 1: Common gene mutations found in E.coli strains

Gene(s) Description Functional Consequence
dam DNA adenine methylase mutation (GATC) Preparing unmethylated DNA, important when trying to cut with certain restriction enzymes (ex: ClaI or XbaI)
dcm DNA cytosine methylase mutation (CCWGG) Preparing unmethylated DNA, important when trying to cut with certain restriction enzymes that are methylation sensitive.
dnaJ Mutation in a chaperonin gene Increases the stability of certain expressed proteins
endA, endA1 Endonuclease I (nonspecific cleavage of dsDNA ) mutation Improves plasmid yield
F Host does (F') or does not (F-) contain the fertility plasmid. A low copy-number plasmid, encodes proteins needed for bacterial conjugation. Genes listed on F´ are wild-type unless indicated otherwise
fhuA (formerly tonA) ferric hydroxamate uptake, iron uptake receptor mutation. T1/T5 Phage resistance
gal Mutation in galactose metabolism pathway Cells cannot grow on galactose only
gyrA, gyrA96 DNA gyrase mutation Confers resistance to nalidixic acid
 hsdRMS hsdR(rk-, mk+) Unmethylated DNA not degraded, cell still can methylate DNA
  hsdS(rk-,mk-) Unmethylated DNA not degraded, cell cannot methylate DNA
lac Lac operon mutations Blue/white screening of clones
  lacIq lac repressor overproduced, expression from pLac repressed more
  LacZ β-galactosidase activity abolished
  lacY Lactose permease inactivated, lactose cannot be taken up by cell
mcrA, mcrBC Inactivation of pathway that cleaves methylated cytosine DNA Allows for uptake of foreign (methylated) DNA
mrr, Δ(mcrC-mrr) Inactivation of pathway that cleaves methylated adenine or cytosine DNA Allows for uptake of foreign (methylated) DNA
recA, recA1, recA13 Mutation in a DNA-dependent ATPase that is essential for recombination and general DNA repair Reduces plasmid recombination, increases plasmid stability
recBCD Exonuclease V activity abolished Increased plasmid stability, reduced recombination
relA or relA1 Relaxed phenotype, mutation eliminating stringent factor Allows RNA synthesis in absence of protein synthesis
Ptrc-ccdA   Propagation of ccdB-containing plasmids
Hte   High transformation efficiency
deoR constitutive expression of genes for deoxyribose synthesis Allows uptake of large plasmids
hee   High electroporation efficiency
supE44 (glnV44)   Suppression of the amber (UAG) stop codon by glutamine insertion
supF (tyrT)   Suppression of the amber (UAG) stop codon by tyrosine insertion
λ-thi-1 or thi1 Mutation in thiamine metabolism Requires exogenous thiamine for growth
ara disruption of arabinose metabolism pathway Inability to utilize arabinose as a carbon source
leuB β-isopropyl malate dehydrogenase inactivated Requires exogenous leucine source for growth
proAB mutation in proline biosynthesis pathway Requires exogenous proline source for growth
rpsL Mutation in subunit S12 of 30S ribosome Confers resistance to streptomycin
Tn10   Confers resistance to tetracycline

Additionally, Table 2 provides a quick reference for some of the popular strains, their genotypes, and their primary use in the lab. These strains are all based on E. coli K-12 and are considered BSL-1.

Table 2: Lab strains of E. coli

Strain Natural resistance Primary Use Genotype
ccdB Survival 2 T1R   TOP10 derivative. For propagating plasmids expressing the ccdB gene (important in Gateway cloning). F-mcrA Δ(mrr-hsdRMS-mcrBC) Φ80lacZΔM15 ΔlacX74 recA1 araΔ139 Δ(ara-leu)7697 galU galK rpsL (StrR) endA1 nupG fhuA::IS2
DB3.1 Streptomycin  HB101 derivative. For propagating plasmids expressing the ccdB gene (important in Gateway cloning). F- gyrA462 endA1 glnV44 Δ(sr1-recA) mcrB mrr hsdS20(rB-, mB-) ara14 galK2 lacY1 proA2 rpsL20(Smr) xyl5 Δleu mtl1
DH10B Streptomycin MC1061 derivative. General cloning and storage, blue/white screening, leucine auxotroph. F- endA1 recA1 galE15 galK16 nupG rpsL ΔlacX74 Φ80lacZΔM15 araD139 Δ(ara,leu)7697 mcrA Δ(mrr-hsdRMS-mcrBC) λ-
DH5alpha   General cloning and storage of common plasmids, blue/white screening. F- endA1 glnV44 thi-1 recA1 relA1 gyrA96 deoR nupG Φ80dlacZΔM15 Δ(lacZYA-argF)U169, hsdR17(rK- mK+), λ–
HB101 Streptomycin Hybrid of E. coli K12 and E. coli B (mostly K12, though), common lab strain for cloning and storage of pBR322 plasmids.  F- mcrB mrr hsdS20(rB- mB-) recA13 leuB6 ara-14 proA2 lacY1 galK2 xyl-5 mtl-1 rpsL20(SmR) glnV44 λ-
JM109   General cloning and plasmid maintenance, blue/white screening, partly restriction-deficient; good strain for cloning repetitive DNA. endA1 glnV44 thi-1 relA1 gyrA96 recA1 mcrB+ Δ(lac-proAB) e14- [F' traD36 proAB+ lacIq lacZΔM15] hsdR17(rK-mK+)
JM110  Streptomycin  For storing plasmids that should not be Dam or Dcm methylated, allows for methylation sensitive restriction enzymes to cut the plasmid after preparation. rpsL thr leu thi lacY galK galT ara tonA tsx dam dcm glnV44 Δ(lac-proAB) e14- [F' traD36 proAB+ lacIq lacZΔM15] hsdR17(rK-mK+) 
MC1061 Streptomycin Parent of DH10B/TOP10 and derived strains, common lab cloning and storage strain. F- Δ(ara-leu)7697 [araD139]B/r Δ(codB-lacI)3 galK16 galE15 λ- e14- mcrA0 relA1 rpsL150(strR) spoT1 mcrB1 hsdR2(r-m+) 
MG1655    Wild type K-12 strain; first published sequence of an E. coli K-12 strain. F- λ- ilvG- rfb-50 rph-1
NEB Stable   For cloning into and storage of lentiviral and retroviral vectors or cloning or repeated sequences with the potential to recombine.  F' proA+B+ lacIq ∆(lacZ)M15 zzf::Tn10 (TetR) ∆(ara-leu) 7697 araD139 fhuA ∆lacX74 galK16 galE15 e14-  Φ80dlacZ∆M15 recA1 relA1 endA1 nupG rpsL (StrR) rph spoT1 ∆(mrr-hsdRMS-mcrBC)
Pir1    For cloning and maintenance of a plasmids with R6Kγ origin; contains a mutant allele of the pir gene that maintains the plasmids at ~250 copies per cell.  F- ∆lac169 rpoS(Am) robA1 creC510 hsdR514 endA recA1 uidA(∆MluI)::pir-116 
Stbl2   JM109-derived. For cloning into and storage of lentiviral and retroviral vectors or cloning or repeated sequences with the potential to recombine.  F- endA1 glnV44 thi-1 recA1 gyrA96 relA1 Δ(lac-proAB) mcrA Δ(mcrBC-hsdRMS-mrr) λ- 
Stbl3  Streptomycin Derived from HB101. For cloning into and storage of lentiviral and retroviral vectors or cloning or repeated sequences with the potential to recombine. This strain is endA+, so column-based DNA preps require an additional wash step. F- glnV44 recA13 mcrB mrr hsdS20(rB-, mB-) ara-14 galK2 lacY1 proA2 rpsL20 xyl-5 leu mtl-1
Stbl4   Electrocompetent cells for cloning and storage of unstable DNA and lentiviral/retroviral vectors. Used by Addgene for pooled library amplification. F' mcrA Δ(mcrBC-hsdRMS-mrr) recA1 endA1 gyrA96 gal-thi-1 supE44 λ-relA1 Δ(lac-proAB)/F' proAB+lacIqZΔM15 Tn10 (TetR)
TOP10 Streptomycin MC1061 derivative. General cloning and storage, blue/white screening. F- mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 nupG recA1 araD139 Δ(ara-leu)7697 galE15 galK16 rpsL(StrR) endA1 λ-
XL1 Blue Tetracycline Blue/white screening and routine cloning, nalidixic acid resistant. endA1 gyrA96(nalR) thi-1 recA1 relA1 lac glnV44 F'[ ::Tn10 proAB+ lacIq Δ(lacZ)M15] hsdR17(rK- mK+) 
XL10 Gold Tetracycline and Chloramphenicol High competency cloning and propagation of large plasmids, ligated DNA, and libraries, nalidixic acid resistant.  endA1 glnV44 recA1 thi-1 gyrA96 relA1 lac Hte Δ(mcrA)183 Δ(mcrCB-hsdSMR-mrr)173 tetR F'[proAB lacIqZΔM15 Tn10(TetR Amy CmR)] 

Note: Inactivating mutations in specific genes are signified with a minus sign (-) as is typically standard; just having the gene listed indicates it is non-functional. If a gene is deleted, it is normally noted with a Greek delta (Δ). 

Additional resources

We've provided an overview of the common lab strains; however, these tables are by no means exhaustive! For a more comprehensive list and additional information, visit OpenWetWare's E.coli genotype resource. Additionally, NEB has a great list of genetic markers for your reference.

Browse Addgene's curated list of Bacterial Expression Systems.

Check out our companion post describing protein expression strains, where we go over the basics of protein expression in E. coli and the common features found in those bacteria.

Check our our blog post covering Ecoli and others are model organisms!

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Topics: Plasmids 101, Plasmids

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