DH5α Escherichia coli Strains
Product Overview: DH5α is the gold standard for molecular cloning, engineered for high transformation efficiency and plasmid stability. Featuring endA1 and recA1 mutations, it produces high-yield, high-purity plasmid DNA while preventing unwanted recombination of inserted fragments. The lacZΔM15 mutation facilitates seamless blue/white colony screening. Optimized for general molecular biology, this strain ensures reliable, reproducible results in cloning, library construction, and routine DNA propagation. Its robust growth characteristics and genetic integrity make it an essential tool for all research laboratories.
Pricing & Availability
| Catalog No. | Packing | Storage | Expiry |
| STR0002 | 100ul | -80℃ | Refer to label |
Technical Specifications
| Genotype | F- endA1 glnV44 thi-1 recA1 relA1 gyrA96 deoR nupG Φ80dlacZΔM15 Δ(lacZYA-argF) U169, hsdR17(rK-mK+), λ- |
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| Resistance | None |
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| Strain Category | E. coli (Competent Cells) |
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| Growth Conditions | LB, 37℃ aerobic |
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Description
The DH5α strain is the most commonly used competent cells in laboratories. The absence of endonuclease (endA) enhances the yield and quality of plasmid DNA; the recombination-deficient (recA) phenotype reduces the probability of homologous recombination of inserted fragments, ensuring the stability of inserted DNA; the presence of lacZΔM15 allows DH5α to be used for blue/white screening. This product is a second-generation glycerol stock (preserved after two generations of amplification from the primary seed), which has not undergone extensive propagation, ensuring stable nuclear DNA and phenotype, with colony phenotype and growth curve matching the original strain by over 99.5%.
Operation Method
Upon receiving the strain, customers should not directly pipette the bacterial suspension for inoculation and propagation. The strain should first be revitalized, and a single colony should be picked for inoculation and propagation after single colonies appear. The revitalization method is as follows: In a biosafety cabinet, open the lid, use an inoculation loop to pick up a small amount of the glycerol stock, and gently streak it on the surface of an LB agar plate using the cross-streak method (Note: Do not puncture the medium). After sealing the plate, incubate at 37°C for 15-20 hours to obtain single colonies.
Cross-streak Method: The purpose of streaking is to achieve gradient dilution of the bacterial suspension through several discontinuous streaks, ensuring the growth of single colonies. Specific steps are as follows:
- A. Rapid Handling: Glycerol stocks taken from -80°C should be opened in a biosafety cabinet, and bacterial liquid/clumps should be immediately picked from the surface for streaking. Do not wait for it to thaw; streaking can be done while it is still solid. The key points are "quick operation, immediate streaking," aiming to minimize "cold shock" and physical damage from ice crystals. If liquid stock is received, streak directly.
- B. Five-Section Method: Figure 1 shows the standard five-section cross-streak method, divided into five zones, with three straight lines streaked in each zone. Zones 1 and 2 can share one inoculation loop. After streaking zones 1 and 2, use a new inoculation loop for zone 3, another new loop for zone 4, and another new loop for zone 5. Each time a new loop is used, it should pass through the streaking marks of the previous zones to effectively dilute the bacterial culture.
- C. Sterilization: Customers may also use a metal inoculation loop, sterilized by flaming, for streaking.
Caution & Storage Guidelines
- 1. This product is FOR RESEARCH USE ONLY!
- 2. Please kindly culture it as soon as possible then harvest and store at -80℃
- 3. Upon receiving the second-generation glycerol stock, it should be immediately stored at -80°C and should not be stored at -20°C.
- 4. The number of freeze-thaw cycles for the glycerol stock should be minimized, as more cycles reduce the viability of the strain.
- 5. If propagating the strain, ensure that no more than four generations are propagated at once to maintain the stability of the strain's genome. After propagation, re-preserved strains must undergo phenotypic and functional validation. Only strains with all phenotypes and functions intact can be used as seed stocks.
Frequently Asked Questions (FAQ)
Q: Why is it necessary to perform strain revitalization upon receipt?
A: Revitalization ensures that you are working with healthy, single-colony isolates. Direct pipetting from frozen stocks can lead to uneven bacterial recovery, reduced transformation efficiency, and potential contamination, which could compromise your downstream experiments.
Q: Why must I store the strain at -80°C instead of -20°C?
A: Storage at -20°C allows for temperature fluctuations and ice crystal formation that severely damage bacterial cell walls and reduce viability. Long-term genetic stability and viability are only guaranteed at -80°C.
Q: What is the significance of the endA1 and recA1 mutations?
A: The endA1 mutation eliminates non-specific endonuclease activity, which prevents the degradation of plasmid DNA during extraction. The recA1 mutation prevents homologous recombination, protecting your inserted DNA fragments from rearrangements or deletions.
Citations & References
(Editor's Note: Please update the citations below to reflect relevant literature.)
- [1] (Example): Hanahan, D. (1983). Studies on transformation of Escherichia coli with plasmids.
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