????DH5α λpir E.coli strain

Description

E.coli DH5αλpir strain is a highly transformablestrain used as a host for suicide plasmid vector.
E. coliDH5αλpir strain contains the pir gene (lysogenized lpir phage lysogen) and isdesigned for cloning and propagation of plasmids with R6K origin of replicationsequence (like pUTmini-Tn5 vectors). To transfer this plasmids directly fromDH5α λpir donor strain to the recipient strain it is necessary to perform atriparental mating with a helper strain such asE. coliDH5α(pRK2013) (see HelperE. colistrains).

Genotype

Thegenotype of the strain is sup E44, ΔlacU169 (ΦlacZΔM15), recA1, endA1, hsdR17,thi-1, gyrA96, relA1, λpir phage lysogen.

Recovery

1.Obtainan LB agar plate with the appropriate antibiotic.

2.Usinga sterile pipette tip, touch the bacteria growing within the punctured area ofthe stab culture. (A sterilized wire loop or sterile toothpick can be used inplace of a sterile pipette tip.)

3.Runthis tip lightly over a section of the plate, as shown in the figure, to createstreak #1.

4.Usinganother sterile pipette tip, pass through streak #1 and spread the bacteriaover a second section of the plate, to create streak #2.

5.Usinga third sterile pipette tip, pass through streak #2 and spread the bacteriaover the last section of the plate, to create streak #3.

6.Growovernight in a 37 ^o C incubator (unless a different growthtemperature is indicated on the plasmid datasheet).

7.Inthe morning, single colonies should be visible. If the bacterial growth is toodense, re-streak onto a new agar plate to obtain single colonies.

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