Question

Hi, I like to make sure I am staying on track with the thought process and was curious if anyone can help with A-D. Thank you so much for the help!

7. (12 pts) You have two Hfr strains of E. coli that carry the F plasmid integrated at different places in the chromosome. During conjugation, strain HfrA transfers the leucine biosynthetic operon only after the rest of the E. coli chromosome has been transferred. During conjugation, strain HfrB transfers the leucine biosynthetic operon early, before any other chromosomal genes get transferred. A third strain, recipient strain C is F (F plasmid free), recA (recA null allele), leu (leucine auxotroph due to null mutation in leu biosynthetic gene) and resistant to streptomycin (Str). Both Hfr strains are streptomycin-sensitive (Str) a) You wish to isolate an F' (F prime) strain that carries the leucine biosynthetic operon. What strains would you use & why? b) What selective medium would you use to select for your strain that carries the F' leu? o) What are the counter selections used that select against donor and recipient strains? d) What difference would it make if your recipient were recA'?

Answer 1

A. To isolate an F' strain carrying leucine biosynthetic operon we would use the HFrB strain which transfers the leucine operon first to ensure the presence of the operon on the F' because if we choose the HfrA strain which sends the leucine operon in the last we might not obtain the gene for the operon because some part in the last of genome is reatined or unable to get transferred to the host.

B. To select the F' strain with leu+ we would grow them in a medium lacking leucine to detect whether they are able to synthesise luecine by their own to survive in the leucine lacking medium or not after the conjugation.

C. To distinguish the donor and recepient we would treat them in medium containing streptomycin antibiotic in which only the donors will die i.e. the Hfr strains because they are streptomycin sensitive and the F- cells will survive which are streptomycin resistant.

D. If the recepints were RecA+ then they will not allow the functioning of the received plasmid to synthesize leucine.