Cosmids vector

1. Dr. Manikandan Kathirvel M.Sc., Ph.D., (NET)
Assistant Professor,
Department of Life Sciences,
Kristu Jayanti College (Autonomous),
Reaccredited with "A++" Grade by NAAC
K. Narayanapura, Kothanur (PO)
Bengaluru
Cosmids

2. Cosmid
Definition
A cosmid is a plasmid that contain phage sequence that allows the vector to be
packaged and transmitted to bacteria like phage vector.
Or
• A cosmid is a type of hybrid plasmid that contains a Lambda phage cos
sequence. Cosmids (cos sites + plasmid = cosmids) DNA sequences are
originally from the lambda phage.

3. Cosmids
1. Cosmids are plasmids, medium sized cloning vectors.
2. Cosmid vector are developed by combining the features of plasmid vector and
bacteriophage vector.
3. The first cosmid vector was described by Collins in 1978.
4. They are capable of incorporating the bacteriophage λ DNA segment. This
DNA segment contains cohesive terminal sites (cos sites).
5. Cos sites are necessary for efficient packaging of DNA into λ phage particles.
6. Large DNA fragments of size varying from 35 to 45 kb can be cloned.
7. They are also packaged into λ. This permits the foreign DNA fragment or genes to
be introduced into the host organism by the mechanism of transduction.
Advantages of using cosmids as vectors:
i. They have high transformation efficiency and are capable of producing a large
number of clones from a small quantity of DNA.
ii. Also, they can carry up to 45 kb of insert compared to 25 kb carried by plasmids
and λ.
Disadvantages of using cosmids as vectors:
i. Cosmids cannot accept more than 50 kb of the insert.

4. Properties of cosmid vector
Contains Features of plasmid
• Origin of replication
• Multiple cloning site
• Selectable marker
Contains Features of lambda phage
• Only cohesive site or cos site region.
• A linear DNA molecule, has a 12 base long , single stranded complementary
overhang at both ends. Which emerge during the packaging process into phage
particle through splitting of cos site.
•• A cosmid vector may have one or two cos site.
Uses:
•• Cosmid vector are used in construction of genomic libraries.
•• Cosmid vector have cloning capacity up to 45 kbp.

5. Cloning of foreign DNA in cosmid vector
involves the following steps:
1. The cosmid is opened at its unique restriction
site and new DNA fragments inserted -Ligation
of foreign DNA.
2. Making a concatemeric DNA.
3. In vitro packaging to introduce the DNA into
phage head to form the matured phage particle.
4. Introduction of the cloned DNA into E. coli by
transduction.
5. After their entry into host cell the cosmid are
maintained as plasmid.
A concatemer is a long continuous DNA molecule that contains multiple copies of the same DNA sequence linked in series.
Examples of Cosmid Vector

6. Cosmid pJB8
Cloning of foreign DNA in cosmid
vector involves the following steps:
1. The cosmid is opened at its unique
restriction site and new DNA
fragments inserted -Ligation of
foreign DNA.
2. Making a concatemeric DNA.
3. In vitro packaging to introduce the
DNA into phage head to form the
matured phage particle.
4. Introduction of the cloned DNA into
E. coli by transduction.
5. After their entry into host cell the
cosmid are maintained as plasmid.

7. A cloning experiment with a cosmid is carried out as follows.
• The cosmid is opened at its unique restriction site and new DNA fragments inserted.
• These fragments are usually produced by partial digestion with a restriction
endonuclease, as total digestion almost invariably results in fragments that are too
small to be cloned with a cosmid.
• Ligation is carried out so that catenanes are formed. Providing the inserted DNA is
the right size, in vitro packaging cleaves the cossites and places the recombinant
cosmids in mature phage particles.
• These lambda phage are then used to infect an E. coli culture, though of course
plaques are not formed.
• Instead, infected cells are plated onto a selective medium and antibiotic-resistant
colonies are grown. All colonies are recombinants, as non-recombinant linear
cosmids are too small to be packaged into lambda heads.