Question

You have three bacterial strains: one is resistant to kanamycin (strain A), another is resistant to gentamycin (strain B), and a third is not resistant to either antibiotic, but produces a purple pigment (strain C). (A) Suppose the antibiotic resistance of one strain is due to a gene on the chromosome, whereas the resistance of the other is due to a gene on a conjugative plasmid. Describe how you could determine which antibiotic resistance is plasmid encoded. (B) Describe how you could determine which, if any, of the strains are naturally competent (i.e., able to take up DNA by transformation).

Answer 1

if we know that one strain has conjugative plasmid and other have chromososomal inheritence for antibiotic resistance, then to determine which antibiotic is plasmid encoded we should perform a conjugation cross with a rec A deficient , antibiotic sensitive strain.As recA protein is essential for recombination so after conjugation we will be sure that no recombination takes place betwwen plasmid DNA and chromosomal DNA. Now if Hfr transfer even takes place the antibiotic resistance gene can't recombine with chromosome hence it will not replicate and lost from the population.But if the antibiotic resistance gene is in the plasmid , they are self replicating so they will be stable in the population. we can also perform antibiotic sensitivity by antibiotic sensitivity test, or plate containing desired antibiotic.

to know if some of the strains are naturally competent then we grow them on CaCl₂ containing plates and incubate them with both heat killed antibiotic resistant bacteria. now we will check for antibiotic sensitivity , if A/B strain found for both antibiotic ressistant then  strain A or B is naturally competent, if C is found antibiotic resistant then Strain C is naturally competent.