Cloning Vectors

At easy level treat cloning vector as a DNA carrier with ori marker and cloning site. Questions are built from these functions.

Ori is for replication marker is for selection and cloning site is for insertion. This is the most useful comparison.

Ori helps in replication but marker is needed for selection. Without marker identifying transformed cells is difficult.

The ori (origin of replication) is the DNA sequence from which vector replication begins. Without an ori, the vector cannot replicate autonomously in the host cell and will not be stably maintained as cells divide. A selectable marker may help in selection, but it cannot compensate for the absence of ori. Exam tip: remember that ori is required for replication, whereas a selectable marker is used to identify or select transformed cells.

MCS provides many designed restriction sites. This allows the insert to be added at a controlled position.

Growth on antibiotic can indicate a transformed cell. But screening may be needed to confirm recombinant status.

Antibiotic selection chooses vector-containing cells. Blue-white screening helps identify insert-containing recombinants.

A blue colony can indicate active lacZ. It generally shows a non-recombinant vector without insert.

A white colony can indicate a recombinant clone due to lacZ disruption. Remember it as a screening result.

ampR gene gives ampicillin resistance. Thus vector-containing cells can grow on an ampicillin plate.

An insert can disrupt a marker gene. This can identify recombinants by insertional inactivation.

Very large vectors can be difficult to introduce into a host. Therefore small size is a common vector feature.

A high-copy-number plasmid is maintained in many copies per host cell, provided that the vector and host system support such maintenance. This feature is useful when the experiment requires a larger yield of plasmid DNA, for example for restriction analysis, sequencing, further cloning, or preparation of a DNA sample. Therefore option A is correct. High copy number does not mean that DNA is removed, that cells are destroyed, or that proteins are digested. The governing trade-off is that increasing copy number can improve DNA yield but may also place a metabolic burden on the host or reduce stability for some inserts. Thus copy number is selected according to the purpose of the cloning experiment.

The direct answer is A, YAC. A very large DNA fragment needs a carrier that can hold much more DNA than an ordinary small plasmid. YAC means yeast artificial chromosome. It contains chromosome-like features that allow large foreign DNA fragments to be maintained in yeast, so it is suitable for very large genomic inserts. Option A works because YACs were designed for large DNA cloning. Option B, a small pUC plasmid, is useful for ordinary smaller inserts but has a much lower carrying capacity, so it is not the best choice here. Option C, pepsin, is a protein-digesting enzyme, not a DNA vector. Option D, amylase, digests starch and is also not a vector. Exam cue: artificial chromosomes are chosen when the insert is too large for a normal plasmid.

BAC stands for bacterial artificial chromosome. It is a bacteria-based cloning vector used to clone relatively large DNA fragments. YAC stands for yeast artificial chromosome, so it is the closest but incorrect option. MCS is a short region containing cloning sites, whereas lacZ is a reporter gene. Exam tip: Link BAC with bacteria and YAC with yeast to remember the abbreviations.

YAC stands for yeast artificial chromosome. It is a cloning vector designed to function like a chromosome in yeast cells and can carry very large DNA fragments. BAC is also used for cloning large DNA fragments, but it stands for bacterial artificial chromosome. Exam tip: Associate YAC with yeast and BAC with bacteria.

Ti plasmid is found in Agrobacterium tumefaciens. Modified Ti plasmid is useful in plant transformation.

The T-DNA of Ti plasmid is transferred into plant cells. In biotechnology it is used after modification.

A bacteriophage is a virus that infects bacteria. Modified bacteriophage vectors can be useful in cloning.

A cosmid carries cos (cohesive-end) sites derived from bacteriophage lambda. These sites are required for packaging DNA into lambda phage particles, so lambda phage DNA packaging is correct. The origin of replication is a separate plasmid sequence and is not the function of a cos site. Exam tip: remember that cosmids combine useful features of plasmids and lambda phage.

The direct answer is A: a plasmid vector is usually a small, circular, double-stranded DNA molecule. To understand this, a plasmid is an extra DNA molecule found independently of the main bacterial chromosome. It can often replicate because it has an origin of replication, and a vector may also contain a selectable marker and cloning site. Option A is correct because it describes the basic physical form of most plasmid vectors. Option B is wrong because a protein chain is not DNA and cannot itself be called a plasmid. Option C is wrong because a fat droplet is a lipid structure, not genetic material. Option D is wrong because chlorophyll is a pigment involved in photosynthesis, not the structure of a vector. Some special plasmids may differ in details, but the basic school-level description remains circular double-stranded DNA. Memory cue: plasmid equals small extra ring of DNA.

Restriction enzymes can prepare ends but DNA ligase performs joining. It makes the recombinant vector.

Directional cloning uses different ends to help insert DNA in the desired direction. It can be useful in expression experiments.

A simple cloning vector may be enough for DNA copy production. Expression may need extra elements like a promoter.

Restriction digestion can indicate presence and size of insert in a vector. Gel pattern helps in confirmation.