This document discusses the Cauliflower Mosaic Virus (CaMV) and its potential use for gene transfer in plants. CaMV is a plant virus that infects brassica plants like cauliflower and turnips. It has a circular double-stranded DNA genome and is spherical in shape. The 35S promoter from CaMV is commonly used in plant transformation due to its strong constitutive expression in dicots. For gene transfer, foreign DNA can be inserted into the non-essential genes II or VII of CaMV. However, CaMV has limitations for gene transfer due to its limited insertion capacity and loss of infectivity if too many nucleotides are added.
2. Plant Viruses:
Plant viruses are considered as efficient gene
transfer agents as they infect the intact plants and
amplify the transferred genes through viral genome
replication
They are non integrative vectors Eg, Pepper mint
mottle virus, Leaf curl virus
3. Criteria needed:
The virus must be capable of spreading from cell to
cell through plasmodesmata
The viral genome should be able to replicate in the
absence of viral coat protein and spreads from cell to
cell
Elicit little or no disease symptoms
Should have broad range of host
4. CaulimoViruses
Caulimoviruses contain circular ds DNA and are
spherical in shape
This group includes 15 viruses of which CaMV is the
most important for gene transfer
The other caulimoviruses includes carnation etched
virus, dahlia mosaic virus mirabilis mosaic virus
and strawberry vein banding virus
5.
6. Cauliflower Mosaic Virus
CaMV is a plant virus that infects mostly brassicaceae
family( cauliflower and turnips) and Solanaceae
species
CaMV is transmitted in a non circulatory manner by
aphid species Mysus
It is a icosahedron structure with a diameter of 52nm
built from 420 cap protein subunits
7. Genome of CaMV
Movement Protein, Insect transmission Factor, Structural
Protein, Translational factor, Reverse Transcription, Cap
8. Thepromoter of the 35S RNA is a very strong constitutive
promoter responsible for the transcription of the whole
CaMV genome.
It is well known for its use in plant transformation.
It causes high levels of gene expression in dicot plants
The 35S RNA is particularly complex, containing a highly
structured 600 nucleotide long leader sequence with six to
eight short open reading frames
This leader is followed by seven tightly arranged, longer
ORFs that encode all the viral proteins. The mechanism of
expression of these proteins is unique, in that the ORF VI
protein (encoded by the 19S RNA) controls translation
reinitiation of major open reading frames on the
polycistronic 35S RNA, a process that normally only
happens on bacterial mRNAs
9. Use of CaMV in gene transfer
For effective transmission of CaMV the foreign DNA must
be encapsulated in viral protein and the inserted gene
should not interfere with native assembly of virus
CaMV does not contain any non coding region wherein
foreign DNA can be inserted
But there a are two genes namely gene II and gene VII have
no essential functions for the virus
So the gene of interest can be replaced in the place of these
two genes
Attractive feature is infection is systemic and the genome
has 3 discontinuities
10. Use of CaMV as a vector pertains to the bacterial dhfr
(dihydroxy folatereductase) gene inserted in the place
of gene II and successfully expressed in plants
This dhfr gene is needed for providing resistance to
methotrexate( inhibitor of dhfr and extremely toxic to
plants)
Limitation of CaMV:
Limited capacity for insertion
Infective capacity is lost if few hundreds of nucleotides
are introduced
Because of its exceeding natural genome size they are
not effectively suitable for gene transfer