Question

what are the main steps in transformation of e. coli? Outline the procedures for each step.

Answer 1

ANSWER: The steps of transformation in bacteria includes-

1.) Choosing a bacterial host- Here we choose E.coli. E.coli is a well studied bacteria with no nuclear membrane and is normally not pathogenic. Also it has enzymes for replication and grows off rapidly.

2.) Choosing a plasmid to transform- An appropriate plasmid have following features :

a.) Single recognition site

b.) origin of replication site

c.) some marker genes like gene for antibiotic resistance

3.) Preparation of E.coli for transformation- for preparation, following steps are followed:

a.) At first, treat with calcium chloride

b.) Electoporation

c.) Lastly inject plasmid into the E.coli cell.

4.) Plate transformation on appropriate media- To choose transformed from non transformed E.coli cells

5.) Incubate the plates overnight- E.coli grows at normal human body temperature( 37 deg). After incubation, analyse the plates.