Question

After transformation, the transformed mixture is plated on 3 different agar plates i). LB, ii). LB/AMP and iii). LB/AMP/ARA. Explain why a bacterial lawn (1000 of bacterial colonies) was observed in i). LB and only few colonies were observed in ii). LB/AMP and iii). LB/AMP/ARA plates?

Answer 1

Ans:- The reason for the 1000 colonies on LB plate after transformation is because the colonies that didn't take up the plasmid will also grow on plate. Lets understand the answer with an example. Suppose, we have the plasmid contains ampicillin metabolizing gene and when this plasmid is transformed and plated onto LB plate and incubated. Then the bacteria that was transformed with plasmid will grow along with the bacterial colony that don't have taken up the plasmid will also grow because there is no antibiotic selection pressure. So, the 1000 of colonies appears in LB plate as transformed as well as untransformed bacteria both grows on the LB plate.

The reason for less or a few colonies in case of transformed plasmid plated for both LB/AMP and LB/AMP/ARA plate is same because the only those colony that are transformed with the plasmid will be able to grow onto ampicillin plate as they contains ampicillin metabolizing gene on the plasmid .Further, non transformed bacterial colony will not grow as they are unable to metabolize the ampicillin present in the plate and die. Thus, growing the transformed colonies onto ampicillin plate keep a selective pressure that only promotes the growth of plasmid containing bacterial cells and untransformed cells die, leads to apperances on only a few colonies onto LB/AMP and LB/AMP/ARA.

Dear student if you find the answer convincing please do give the feedback.