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Cloning Vectors

Cloning vectors are DNA molecules that carry foreign DNA into host cells and replicate to produce multiple copies. They contain sequences for propagation in bacteria or yeast, a cloning site for inserting foreign DNA, and a selectable marker for identifying cells containing the vector. Common vectors include plasmids, phages, cosmids, BACs, and YACs, which have different cloning limits ranging from a few kilobases to over a megabase. The general cloning process involves digesting the vector and DNA with restriction enzymes, ligating the foreign DNA into the vector, transforming the DNA into host cells, and selecting cells containing the foreign DNA.

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220 views1 page

Cloning Vectors

Cloning vectors are DNA molecules that carry foreign DNA into host cells and replicate to produce multiple copies. They contain sequences for propagation in bacteria or yeast, a cloning site for inserting foreign DNA, and a selectable marker for identifying cells containing the vector. Common vectors include plasmids, phages, cosmids, BACs, and YACs, which have different cloning limits ranging from a few kilobases to over a megabase. The general cloning process involves digesting the vector and DNA with restriction enzymes, ligating the foreign DNA into the vector, transforming the DNA into host cells, and selecting cells containing the foreign DNA.

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Raihanur Kiran
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Cloning Vectors

The molecular analysis of DNA has been made possible by the cloning of DNA. The two
molecules that are required for cloning are the DNA to be cloned and a cloning vector.

Cloning vector - a DNA molecule that carries foreign DNA into a host cell, replicates inside a
bacterial (or yeast) cell and produces many copies of itself and the foreign DNA.

Three features of all cloning vectors

1. Sequences that permit the propagation of itself in bacteria (or in yeast for YACs)
2. A cloning site to insert foreign DNA; the most versatile vectors contain a site that can be
cut by many restriction enzymes.
3. A method of selecting for bacteria (or yeast for YACs) containing a vector with foreign
DNA; uually accomplished by selectable markers for drug resistance.

Types of Cloning Vectors


Plasmid - an extrachromosomal circular DNA molecule that autonomously replicates
inside the bacterial cell; cloning limit: 100 to 10,000 base pairs or 0.1-10 kilobases (kb)
Phage - derivatives of bacteriophage lambda; linear DNA molecules, whose region can
be replaced with foreign DNA without disrupting its life cycle; cloning limit: 8-20 kb
Cosmids - an extrachromosomal circular DNA molecule that combines features of
plasmids and phage; cloning limit - 35-50 kb
Bacterial Artificial Chromosomes (BAC) - based on bacterial mini-F plasmids. cloning
limit: 75-300 kb
Yeast Artificial Chromosomes (YAC) - an artificial chromosome that contains telomeres,
origin of replication, a yeast centromere, and a selectable marker for identification in
yeast cells; cloning limit: 100-1000 kb

General Steps of Cloning with Any Vector


1. Preparation of the vector and DNA to be cloned by digestion with restriction enzymes to
generate complementary ends
2. Ligation of the foreign DNA into the vector with the enzyme DNA ligase.
3. Introduction of the DNA into bacterial cells (or yeast cells for YACs) by transformation.
4. Selection of cells containing foreign DNA by screening for selectable markers (usually
drug resistance).

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