Question

In the selection of bacterial colonies that carry cloned DNA in plasmids, such as \(\mathrm{pBR} 322\), that contain two antibiotic resistance genes, A. one antibiotic resistance gene is nonfunctional in the desired bacterial colonies. B. untransformed bacteria are antibiotic resistant. C. both antibiotic resistance genes are functional in the desired bacterial colonies. D. radiolabeled DNA or RNA probes play a role. E. none of the above.

Short answer

A. One antibiotic resistance gene is nonfunctional in the desired bacterial colonies B. Untransformed bacteria are antibiotic resistant C. Both antibiotic resistance genes are functional in the desired bacterial colonies D. Radiolabeled DNA or RNA probes play a role E. None of the above Answer: C. Both antibiotic resistance genes are functional in the desired bacterial colonies

Step-by-step solution

Understanding the pBR 322 Plasmid and Cloned DNA

pBR 322 is a widely used plasmid vector in molecular cloning. It is a small, circular piece of DNA that has two antibiotic resistance genes: ampicillin resistance (ampR) and tetracycline resistance (tetR). These resistance genes are crucial for the selection of bacterial colonies that have successfully taken up the recombinant plasmid containing the desired cloned DNA.

Option A: One antibiotic resistance gene is nonfunctional in the desired bacterial colonies

This statement is incorrect because both antibiotic resistance genes are functional in the desired bacterial colonies, ensuring their survival in the presence of both antibiotics.

Option B: Untransformed bacteria are antibiotic resistant

This statement is incorrect. Untransformed bacteria do not carry the antibiotic resistance genes and will not survive in the presence of the antibiotics, ensuring that only transformed bacteria containing the desired cloned DNA will be selected.

Option C: Both antibiotic resistance genes are functional in the desired bacterial colonies

This statement is correct. Both antibiotic resistance genes present in the pBR 322 plasmid are functional in the desired bacterial colonies, ensuring their survival in the presence of both antibiotics. These resistance genes help in the selection of transformed bacteria carrying the desired cloned DNA.

Option D: Radiolabeled DNA or RNA probes play a role

This statement is incorrect. Radiolabeled DNA or RNA probes are not directly involved in the selection of bacterial colonies containing cloned DNA in pBR 322 plasmids. These probes may be used in other techniques for monitoring gene expression or detecting specific sequences within the cloned DNA insert, but they are not critical for the actual selection of transformed bacteria.

Option E: None of the above

This statement is incorrect, as option C correctly describes the role of both antibiotic resistance genes being functional in the desired bacterial colonies.

Conclusion

The correct answer to the multiple-choice question is C. Both antibiotic resistance genes (ampR and tetR) present in the pBR 322 plasmid are functional in the desired bacterial colonies, ensuring their survival in the presence of both antibiotics and helping in the selection of transformed bacteria carrying the desired cloned DNA.

Key concepts

Molecular Cloning

Molecular cloning is a fundamental technique in biosciences that involves the isolation and amplification of a specific DNA sequence. The process begins with the extraction of the desired gene or DNA fragment. This DNA is then inserted into a vector, like the pBR322 plasmid, which will carry the gene. The vector is introduced into a host cell, commonly bacteria, through a process known as transformation.

After transformation, only bacteria that have successfully taken up the plasmid grow in the presence of antibiotics, thanks to the antibiotic resistance genes in the vector. By using specific antibiotics that correspond to the resistance genes in the plasmid, researchers can select for bacteria that have incorporated the plasmid and, consequently, the gene of interest. The simplicity and effectiveness of molecular cloning have made it essential for studying genes and proteins, gene therapy, and the production of recombinant proteins.

Antibiotic Resistance Genes

Antibiotic resistance genes are segments of DNA that confer survival advantage to bacteria exposed to antibiotics. These genes are an integral part of molecular cloning and are used as markers in the selection process. For instance, the pBR322 plasmid holds two important antibiotic resistance genes: one for ampicillin and another for tetracycline.

Selecting Transformed Cells

The presence of these genes in the cloning vector allows scientists to easily identify which cells have been successfully transformed. When a bacterial culture is grown on a medium containing antibiotics, only those bacteria with the plasmid containing the resistance genes will survive. Consequently, this provides a convenient way to select only the bacterial colonies that carry the cloned DNA.

Recombinant DNA Technology

Recombinant DNA (rDNA) technology involves combining DNA molecules from different biological sources to create new genetic sequences with desired functions. The backbone of this technology is the capability to cut and ligate DNA molecules using restriction enzymes and ligases, respectively.

Creating a Plasmid with New Traits

With pBR322 as an example, scientists insert a foreign DNA fragment into the plasmid's cloning site. This recombinant plasmid can then be introduced into host cells, creating a powerful way to study gene function, produce proteins, and develop genetically modified organisms. rDNA technology has paved the way for advancements in medicine, agriculture, and industrial biotechnology.

Bacterial Transformation

Bacterial transformation is the process by which bacteria take up foreign DNA from their surroundings. It is a critical step in various biotechnological procedures, including molecular cloning. Through transformation, the pBR322 plasmid carrying foreign DNA can be introduced into a bacterial host.

Ensuring Successful Transformation

To ensure that transformation has been successful, and that bacteria have indeed incorporated the plasmid DNA, scientists rely on antibiotic resistance genes as markers. Only the bacteria that grow on antibiotic-containing media are considered transformed. This process is essential because it is the starting point for producing multiple copies of the inserted gene, enabling further study and utilization of the gene's products.