Arrange the following steps involved in the transformation of bacteria in a sequence from initiation to end.

(A) Incubation of rDNA with bacterial cell on ice

(B) Treatment with divalent cations
(C) Heat shock treatment
(D) Selection on antibiotic containing agar plate
(E) Placed them again on ice

Choose the correct answer from the options given below:

The correct answer is (B), (A), (C), (E), (D).

Here is the step-by-step educational explanation of the bacterial transformation process:

Step 1: Treatment with divalent cations (B)
Before introducing recombinant DNA (rDNA) into bacterial cells (such as Escherichia coli), the host cells must be made competent to take up foreign DNA. This is achieved by treating the bacterial cells with a specific concentration of a divalent cation, such as calcium (${\mathrm{Ca}}^{2+}$). These cations increase the efficiency with which DNA enters the bacterium through pores in its cell wall.

Step 2: Incubation of rDNA with bacterial cell on ice (A)
Once the cells are made competent, the recombinant DNA (rDNA) is added to the mixture. The recombinant DNA and the competent bacterial cells are incubated together on ice. This allows the DNA to bind to the cell wall of the bacteria.

Step 3: Heat shock treatment (C)
The mixture is then subjected to a brief heat shock, usually by placing it at 42°C for about 30 to 90 seconds. This sudden rise in temperature creates transient thermal pores in the bacterial cell membrane, allowing the rDNA to enter the cell.

Step 4: Placed them again on ice (E)
Immediately following the heat shock, the bacteria are placed back on ice. This rapid cooling helps to close the pores and stabilize the bacterial cell membrane, securing the transformed DNA inside the cell.

Step 5: Selection on antibiotic containing agar plate (D)
Finally, the cells are plated on a selective agar medium containing an antibiotic. Since the vector (plasmid) used to make the rDNA typically carries a gene for antibiotic resistance, only the successfully transformed bacteria (cells that have taken up the rDNA) will survive and grow into colonies on this plate. This step allows for the selection of the transformants.