Question

A scientist used a plasmid that contained ampicillin resistance gene and lacZ gene as selectable markers. He inserted the gene of interest within the ampicillin resistance gene instead of lacZ gene due to human error. The competent cells were allowed to take up the plasmid. This was followed by plating them on agar media containing ampicillin and X-gal. Which of the following statements correctly describes the result of the experiment?

• A. The bacteria which took up the recombinant plasmid would not grow
• B. The bacteria which took up the recombinant plasmid would form white colonies
• C. All of the bacteria would grow and give white colonies
• D. The bacteria which took up the recombinant plasmid would grow and give blue colonies

Answer 1

The correct option is A The bacteria which took up the recombinant plasmid would not grow

The ampicillin resistance gene in vectors provides resistance against the antibiotic, ampicillin, and is used as a selectable marker. In this experiment, the scientist has accidentally inserted the gene of interest within the ampicillin resistance $(Am{p}^R$) gene. This would inactivate the $Am{p}^R$ gene and make all the host cells sensitive to ampicillin. So, the transformed bacteria which took up the plasmids would not be able to grow on agar plate containing ampicillin.

The lacZ gene is used as a selectable marker in many cloning vectors to screen recombinants from non-recombinants. It codes for protein 𝛃-galactosidase which is an enzyme. This enzyme hydrolyses the chromogenic substrate X-gal into a blue product.

In another scenario, if the gene of interest would have been inserted within the lacZ gene as planned, then the ampicillin resistance gene would have remained intact. Also, the 𝛃-galactosidase enzyme would not have been synthesised in the host cell due to insertional inactivation. Thus, X-gal would not have been converted into a blue product. This would have resulted in the production of white colonies.