Top 10 vs DH5α? - TOPO TA cloning question … (Jan/16/2007 )
Can anyone please explain the differences between the Top 10 and the DH5α strains that come with the TOPO TA cloning kit (Promega)?
Not a huge difference. I believe the TOP10 have a higher transformation efficiency.
TOP10 E. coli are provided at a transformation efficiency of 1 x 109 cfu/µg supercoiled DNA and are ideal for high-efficiency cloning and plasmid propagation. They allow stable replication of high-copy number plasmids. The genotype of TOP10 Cells is similar to the DH10B™strain, and offers the following features:
* hsdR for efficient transformation of unmethylated DNA from PCR amplifications
mcrA for efficient transformation of methylated DNA from genomic preparationslacZM15 for blue/white color screening of recombinant clonesendA1 for cleaner preparations of DNA and better results in downstream applications due to the elimination of non-specific digestion by Endonuclease IrecA1 for reduced occurrence of non-specific recombination in cloned DNAOne Shot® TOP10 Kits are available with either chemically competent or electrocompetent E. coli to fit your specific transformation needs. TOP10 E. coli are also available in the high-throughput MultiShot™ format.
F- mcrA (mrr-hsdRMS-mcrBC) 80lacZM15 lacX74 recA1 ara139 (ara-leu)7697 galU galK rpsL (StrR) endA1 nupG
DH5™ is a well-known, versatile strain that can be used in many everyday cloning applications. In addition to supporting blue/white screening, recA1 and endA1 mutations in DH5™ increase insert stability and improve the quality of plasmid DNA prepared from minipreps. DH5™ Cells offer the following benefits:
* A wide range of efficiencies from >1 x 106 to>1 x 109 transformants/µg
* Greatly increased plasmid yield and quality due to endA1 mutation
* Blue/white screening of recombinant clones due to lacZM15
* Ensured insert stability due to recA1 mutation
F- 80dlacZM15 (lacZYA-argF) U169 recA1 endA1 hsdR17(rk-, mk+) phoA supE44 - thi-1 gyrA96 relA1
someone told me that top10 are not good to transform retroviral vectors.. i dont know the principles sorry, but i hear that modification of the plasmid due to recombination could happen
Thanks, that makes more sense now!
There is no difference unless you are creating a library from a methylated eukaryotic DNA source (DH10B lacks some of the restriction systems that prevent unbiased library creation).