Takara E. coli chemically competent cells

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Applications

• Recombinant DNA cloning experiments, such as library preparation, generation of recombinant plasmids, or subcloning
• Site-directed mutagenesis (HST02 and BMH71-18 mutS)
• Cloning methylated DNA (HST08 or HST16CR)
• Cloning large DNA fragments (HST08 or HST16CR)

Purity

No colonies form after inoculation of LB-agar medium containing 100 µg/ml ampicillin with 100 µl of E. coli competent cells.

Genotypes

E. coli HB101 (Cat. # 9051): e14–(mcrA_–), _supE_44, _relA_1, Δ(_lac-proAB)/F` [traD_36, _proAB+, _lac I_q, _lacZ_ΔM15]

E. coli CJ236 (Cat. # 9053): _dut_1, _ung_1, _thi_-1, _relA_1/pCJ105(F` _cam_r)

E. coli BMH71-18 mutS (Cat. # 9054): Δ(lac-proAB), supE, thi_-1, _mutS_215:: Tn_10 (tet_r)/F` [_traD_36, _proAB+, _lac I_q, _lacZ_ΔM15]

E. coli MV1184 (Cat. # 9055): ara, Δ(lac-proAB), rpsL, thi ( ø80_lacZ_ΔM15), Δ(srl-recA)306:: Tn_10_ (tet_r)/ F`[_traD_36, _proAB+, _lac I_q, _lacZ_ΔM15]

E. coli HST02 (Cat. # 9127): F` [tra_D36, _pro_A+B+, _lac_Iq, _lac_ZΔM15]/Δ(_lac_-_pro_AB), _rec_A, _end_A, _gyr_A96, _thi, e14-(_mcr_A-), _sup_E44, _rel_A, Δ_deo_R, Δ(_mrr_-_hsd_RMS-_mcr_BC)

E. coli DH5α (Cat. # 9057): F–, ø80d_lacZ_ΔM15, Δ(lacZYA-argF)U169, deo_R, _recA_1, _endA_1, _hsdR_17(rK–, mK+), _phoA, _supE_44, λ–, _thi_-1, _gyrA_96, _relA_1

E. coli HST08 (Cat. # 9128): F–, endA_1, _supE_44, _thi_-1, _recA_1, _relA_1, _gyrA_96, _phoA, ø80d_lacZ_ΔM15, Δ(lacZYA-argF)U169, Δ(_mrr-hsd_RMS-_mcr_BC), Δ _mcr_A, λ–

E. coli HST16CR (Cat. # 9131): F–, endA_1, _supE_44, _thi_-1, _recA_1, _relA_1, _gyrA_96, _phoA, ø80d_lacZ_ΔM15, Δ(_lacZYA-arg_F)U169, Δ(_mrr-hsd_RMS-_mcr_BC), Δ _mcr_A, Δ _pcn_B, λ–

Uses of each strain

E. coli HB101: HB101 maintains a stable genotype. It is used as a general-purpose strain for recombinant DNA experiments.

E. coli JM109: JM109 is used for alpha-complementation (blue/white screening) in which clones containing recombinant plasmids are identified by screening for beta-galactosidase activity. The F` episome in JM109 includes _lacZ_ΔM15. If a host cell contains a plasmid encoding _lacZ_alpha in the MCS region (such as pUC vectors), the colony will appear blue when plated in the presence of X-gal. If _lacZ_alpha in the MCS region of the plasmid has been disrupted by an insert, the resulting colony will appear white when plated in the presence of X-gal. JM109 can be used to prepare ssDNA as well as for library construction and subcloning.

E. coli CJ236: CJ236 is used for preparation of uracil-containing ssDNA (Kunkels method). CJ236 is _dut_– _ung_– and thereby lacks dUTPase and uracil-DNA-glycosylase, causing mis-incorporation of deoxyuridine in place of some thymidine residues. Kunkels method of site-directed mutagenesis utilizes this strain as the host of pUC118/119 or M13 phage to prepare the template ssDNA. Stable maintenance of the F` episome (pCJ105) requires cultivation in the presence of chloramphenicol.

E. coli BMH71-18 mutS: BMH71-18 _mut_S is used for site-directed mutagenesis, and lacks the ability to repair mismatched DNA. Therefore, this host suppresses repair of mismatched sites and increases the efficiency of mutagenesis.

E. coli MV1184: MV1184 is used to select amber mutants. MV1184 contains the F` episome and can be used as an M13 phage vector or phagemid vector host to prepare ssDNA.

E. coli HST02: HST02 contains the F` episome and can be used as an M13 host, to prepare DNA libraries, or for subcloning. The restriction-minus phenotype (mcrA, mrr-mcr_BC _hsd_RMS) of HST02 enables efficient cloning of methylated DNA. HST02 can be used for alpha-complementation (blue/white screening) in which clones containing recombinant plasmids are identified by screening for beta-galactosidase activity.
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E. coli_ DH5α:** DH5 alpha is the most frequently used E. coli strain for general-purpose cloning. It may be used for alpha-complementation (blue/white) screening in which clones containing recombinant plasmids are identified by screening for beta-galactosidase activity, e.g., in combination with pUC vectors. DH5 alpha does not carry _lacl_q and therefore IPTG is not required when performing blue/white screening, making the screening step more economical.

E. coli HST08: HST08 lacks the genes required for restriction of foreign methylated DNA (mrr, _hsd_RMS, _mcr_BC, and _mcr_A). It is recommended when cloning large inserts greater than 10 kb in combination with the TaKaRA DNA Ligation Kit LONG (Cat. # 6024). HST08 is useful for a wide range of applications including cloning of methylated DNA.

Transformation efficiency

When using 1 ng plasmid DNA to transform 100 µl of cells:

HB101 Competent Cells	>1 × 108 cfu/µg pBR322
JM109 Competent Cells	>1 × 108 cfu/µg pBR322
CJ236 Competent Cells	>1 × 107 cfu/µg pUC119
BMH71-18 mutS Competent Cells	>1 × 107 cfu/µg pUC119
MV1184 Competent Cells	>1 × 107 cfu/µg pUC119
HST02 Competent Cells	>1 × 108 cfu/µg pUC19
DH5α Competent Cells	>1 × 108 cfu/µg pUC19
HST08 Premium Competent Cells	>1 × 108 cfu/µg pUC19
HST16CR Competent Cells	>1 × 108 cfu/µg pUC19

Stability of F` plasmid, confirmation of α-complementation:

• E. coli JM109, E. coli BMH71-18 mutS, E. coli MV1184, E. coli HST02
Less than 1% of colonies with a white phenotype appeared when transformed with pUC19 (for JM109 or HST02) or pUC119 (for BMH71-18 mutS or MV1184) followed by plating on L-agar medium containing 100 µl/ml ampicillin, 0.2 mM IPTG, and 40 µg/ml X-Gal.

• E. coli CJ236
Strains lacking F` plasmid (pCJ105) do not grow on a plate containing 30 µg/ml chloramphenicol.

• E. coli DH5α
Blue colonies appeared when cells were transformed using pUC19 DNA and then plated on L-agar medium containing 100 µg/ml of ampicillin and 60 µg/ml of X-Gal.

Additional product information

Please see the product`s Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.

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