Question

Which of the following vector can be used for obtaining single-strand copies of a cloned sequence?

(1). pUC18
(2). Cosmid
(3). \( \lambda \) phage
(4). Phage M13

• A. pUC18
• B. Cosmid
• C. \( \lambda \) phage
• D. Phage M13

Hint:

Whenever single-stranded DNA vectors are mentioned in biotechnology, think of M13 or phagemid vectors (like pBluescript containing an f1 origin of replication).

Answer 1

The Correct Option is D

Step 1: Understanding the Concept:
Certain applications, such as Sanger dideoxy sequencing, site-directed mutagenesis, and hybridization probes, historically required or benefited from obtaining pure single-stranded DNA (ssDNA) of a cloned gene of interest.

Step 2: Detailed Explanation:
Let us evaluate the nature of each vector:
1. pUC18 (1): This is a standard plasmid vector. It replicates inside the host as double-stranded DNA (dsDNA).
2. Cosmid (2): Cosmids are hybrid vectors combining plasmid features with the cos sites of lambda phage. They are packaged into phage heads but replicate in host cells as large double-stranded plasmids.
3. \( \lambda \) Phage (3): This is a double-stranded DNA bacteriophage used for cloning larger DNA fragments.
4. Phage M13 (4): M13 is a male-specific, filamentous bacteriophage of E. coli that contains a single-stranded circular DNA genome.
- When M13 infects a host, its ssDNA is converted into a double-stranded replicative form (RF), which can be manipulated like a plasmid.
- During the late stage of its life cycle, M13 replicates via a rolling-circle mechanism to produce single-stranded circular genomes.
- These single-stranded genomes are packaged into protein coats and extruded through the host cell wall without lysing the host.
- This allows researchers to easily harvest pure single-stranded viral DNA containing the cloned insert directly from the culture supernatant.

Step 3: Final Answer:
Phage M13 is the vector used to obtain single-strand copies, corresponding to option (D).