Biology:DH5-Alpha Cell
From HandWiki
DH5-Alpha Cells are E. coli cells engineered by American biologist Douglas Hanahan to maximize transformation efficiency. They are defined by three[1] mutations: recA1, endA1 which help plasmid insertion and lacZΔM15 which enables blue white screening. The cells are competent and often used with calcium chloride transformation to insert the desired plasmid. A study of four transformation methods and six bacteria strains showed that the most efficient one was the DH5 strain with the Hanahan method.[2]
Mutations
- The recA1 mutation is a single point mutation that replaces glycine 160 of the recA polypeptide with an aspartic acid residue[3] in order to disable the activity of the recombinases and inactivate homologous recombination.
- The endA1 mutation inactivates an intracellular endonuclease to prevent it from degrading the inserted plasmid.[4]
References
- ↑ "Strain - DH5α". The Coli Genetic Stock Center (CGSC). Yale University. https://cgsc2.biology.yale.edu/Strain.php?ID=150015.
- ↑ "A comparison and optimization of methods and factors affecting the transformation of Escherichia coli". Bioscience Reports 33 (6). December 2013. doi:10.1042/BSR20130098. PMID 24229075.
- ↑ "Construction of a recombinase-deficient mutant recA protein that retains single-stranded DNA-dependent ATPase activity". The Journal of Biological Chemistry 263 (18): 8716–8723. June 1988. doi:10.1016/S0021-9258(18)68364-4. PMID 2967815. http://www.jbc.org/content/263/18/8716.full.pdf.
- ↑ "E. coli host strains significantly affect the quality of small scale plasmid DNA preparations used for sequencing". Nucleic Acids Research 21 (7): 1677–1678. April 1993. doi:10.1093/nar/21.7.1677. PMID 8479929.
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