MOLECULAR SWITCHES, INSTALLMENT #2, MINIMAL GENOME PROJECT

The Minimal Genome Project

JENNIFERLAKE

May 11, 2011 at 10:40 pm

Remember SARS in 2003? Looking back on SARS led me to something dubbed the Minimal
Genome Project which I found a lot of experimental documentation for between 1998 and 2000,
including the creation of completely new amino acids --until this, every biological entity that has
ever existed used the same available 20 amino acids. SARS patients were found to be infected
with a novel corona virus. Coincidently, a novel corona virus was created in a lab with a new
amino acid in 2001. Since the original outbreak, the SARS creature seems to have disappeared . .
.
Chalk this up to spooky things scientists say.

November 22, 2002 -- "Craig Venter's 'minimal genome' project announced Wednesday is not
about creating a new life form...[that comes later]. The high profile project was just funded by
the U.S. Department of Energy (DoE) with $3 million going to the Institute for Biological
Energy Alternatives (IBEA), one of the nonprofit research institutes Venter founded after
leaving..Celera Genomics early this year.

"The question of the minimal genome for an organism [--the base gene set required for life--] is
always 'minimal in which environment,' said Francisco J. Silva of the University of Valencia in
Spain. Silva and colleagues..are studying the genome of the insect endosymbiont Buchnera
aphidicola, which appears to have an even smaller genome than that of the parasite Mycoplasma
genitalium, Venter's organism of choice.

"..pathogenic bacteria usually have more genes than their harmless relatives do..but the disease-
related genes are not essential to life, so they could be the first catagory of genes a scientist
would jettison from a minimal genome organism... 'A minimal genome organism' [Silva] said,
'will be a prisoner of the laboratory dish where it lives, and will be unable to compete with the
outer world.'

"The minimal genome organism now being planned..will be further crippled so that it cannot
survive wihout laboratory coddling. The strategy is to synthesize an artificial chromosome
containing the presumptive minimal gene set, remove all existing genetic material.. and then
insert the synthetic chromosome into the vacant cell... The long-term goal..will be to help the
world solve some of its environmental problems. That's why the funding is coming from DoE,
where recent genetics projects have focused on bioremedation..."

I went searching for the citation in her blog but couldn't find the source. I did come up with:

http://www.gene-watch.org/blog/post/Venter-Speaks-Out-on-the-Human-Genome-Project-
Synthetic-Biology-and-Personal-Genomics-Among-Other-Things.aspx
where the insano Venter put an email address in a genome that people decoded and responded to
to show that clandestine operations desired by the KGB for all these years has finally been
achieved. And you thought the US and USSR were two different entities

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http://www.economist.com/node/3422878?story_id=3422878

Then I finally got the citation for him bioprospecting for lifeforms in the ocean in his
SORCERER boat.

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THEN I FOUND THIS WEB RESOURCE THAT I DATAMINED FOR YOU. THIS
COLLECTION IS LONG. THERE ARE SOME ON MY LIST THAT DO THE HARD
WORK. IF YOU KNOW WHAT IS GOOD FOR YOU AND THE WORK AT HAND THEN
YOU WILL SPEND THE TIME TO READ THE ENTIRE THING.

http://www.molecular-plant-biotechnology.info/genomics-and-bioinformatics/minimum-
genome-size.htm

Minimum Genome Size - The minimum number of genes required to sustain life can at least be
only guessed at. This guess is based on the following. The genes present in different organisms
having the smallest of genomes and their functions are compared critically. In addition,
experimental results are gathered by inactivating specific genes of these organisms and assessing
their effects on the organism's survival.

Based on these analyses, it has been estimated that living organisms require a minimum of 250-
350 genes. This minimum gene, number is necessary for the organisms to exist as independent,
self replicating organisms.

http://www.molecular-plant-biotechnology.info/immune-systems-antibodies-interferons-
vaccines/immune-systems-antibodies-interferons-vaccines.htm

Immune System, Antibodies, Interferons and Vaccines -

Immunology deals with the defense mechanisms of animals against parasites, so that these
animals are normally capable of resisting the infection by most pathogens. However, the
biochemical and molecular basis of such resistance has been understood only during the last few
decades.

The pathogens are always recognized by the host cells through chemical interactions; a specific
antigen or immunogen, present in the parasite's body, elicits in the host the production of a
specific glycoprotein complex (an antibody) or an immunoglobulin (Ig).

The antibody or immunoglobulin is always a protein, covalently bound with an oligosaccharide.

In contrast, although most often, antigen is a protein, it may also be a polysaccharide or nucleic
acid or any other substance.
It is also possible that a foreign protein (not necessarily belonging to a pathogen) may act as an
antigen so that when injected, it may induce antibody formation. Certain low molecular weight
molecules called haptens, ray though bind to antibodies, do not individually stimulate antibody
production.

However, they may become antigenic and stimulate antibody production, if they are tightly
bound to certain macromolecules such as proteins, polysaccharides and nucleic acids.

It should also be recognized that the entire surface of an antigen molecule is not necessary for its
antigenicity; instead specific group of atoms called antigenic determinant or epitope, consisting
of 5-8 amino acids, is needed for immune response and antibody production.

The different areas of Immunotechnology involving, both basic and applied aspects, will be
briefly discussed in this chapter. In the next chapter, genetic engineering and uses of monoclonal
antibodies will he discussed.B

http://www.molecular-plant-biotechnology.info/immune-systems-antibodies-interferons-
vaccines/genetically-engineered-viruses-as-vaccines.htm

Genetically Engineered Viruses as Vaccines -

Although in the past attenuated strains of vaccinia virus were obtained by serial virus passage,
have been used for this purpose in 1980's and 1990's Deletion of several genes (thymidine
kinase, a growth factor, hemagglutinin, 13.8 kD secreted protein, and ribonucleotide reductase)
decreases its virulence.

Similarly insertion of some lymphokine genes (e.g. interleukin-2 or IL-2) also lead to decreased
virulence. While reducing virulence or pathogenicity, immune response of the recombinant
protein (coded by gene inserted in the virus for developing specific vaccine) must be increased.

More than one gene can also be stably integrated into attenuated
vaccinia virus for developing recombinant virus to be used as a vaccine
against multiple pathogens.

A glycoprotein gene of rabies virus has been integrated and the resulting
rabies virus vaccine has been field tested in Europe and U.S.A. giving
promising results.
Other vaccines against HIV are being developed using the gene for HIV-1 envelope
glycoprotein. In future, the widespread use of Vaccinia as a live vaccine will depend on
improving safety, while achieving higher response to the recombinant protein.

THE FORCING OF THE EXPRESSION OF FOREIGN SUBSTANCES FROM THE CELLS

TAKES OUT ALL OF THE ENERGY OF THE CELL METABOLISM WHICH IS THE
DEFINITION OF CHRONIC FATIGUE.
OK, KIDLINGS. THIS ONE NEARLY PUT ME TO SLEEP. I WOULD HAVE SPARED
YOU THE MIDDLE PORTION BECAUSE I READ THE OPENING PARAGRAPH THEN
WENT: WTF? WHEN THEY DID A 180 ON THE UNITED NATIONS CRAP. WHERE DID
THAT COME FROM? BUT THEN THE PAYOFF WAS AT THE END HIGHLIGHTED IN
RED. IF YOU UNDERSTAND THAT THEY WRITE THIS STUFF IN CODE FOR 'THEM'
THE MESSAGE IS BLATANT:
YOU ARE WASTE ON THE PLANET THAT NEEDS BIOREMEDIATION VIA VACCINE!

http://www.molecular-plant-biotechnology.info/immune-systems-antibodies-interferons-
vaccines/nature-of-vaccines.htm

Nature of Vaccines -
In broader sense, the term vaccine is used for any biological material injected in the body to
stimulate the active immunity. Immunization of the body through vaccination is generally done
for the prevention of disease.

The vaccine is administered in advance so as to give the body time to set active immunity, before
invasion by the pathogen occurs. The objective of vaccination is to stimulate the production of
antibodies without the person actually having to develop the disease.

(ii) After 20 yeas, in June 1992, United Nations Conference on Environment and
Development (UNCED), popularly described as Rio Earth Summit, was help in Rio de
Janeiro (Brazil), where heads of the states from 166 countries participated to examine the issues
involved and the solutions possible.
(iii) The latest conference, the World Summit on Sustainable Development (WSSD) was held
in Johannesburg (South Africa) during August 26 to September 4, 2002 to assess global change
since the above Rio Earth Summit.
While on the one hand, there is an increasing problem of the conservation of nature and natural
resources. Both these problems are receiving constant attention of environmentalists. Among
implications, there is also an alarm due to release of genetically engineered organisms in the
atmosphere and due to release of effluents from biotechnological companies, so that the
environmentalists are having a debate on the effects of developments in biotechnology on the
environment.

There is also a debate on the safety of the use of the products of biotechnology, an area described
as biosafety. Among application, on the other hand, efforts are also being made to use
biotechnology to protect the environment from pollution and to conserve natural resources. At a
time, when the gap between those who have plenty and those who do not have even the
minimum is widening, both ends of this spectrum, i.e. plenty and poverty, are contributing to
environmental degradation. It is, therefore, necessary that the developing and developed
countries jointly find a path of development which meets the needs of the present without
compromising the ability, of future generations to meet their needs (World Commission on
Environment and Development). Efforts are being made to achieve this objective through a
variety of approaches, and biotechnology is certainly one of them. In this chapter and the next
four chapters, environmental; implications and applications of biotechnology for environment
will be discussed.
In recent years, we have witnessed a debate on the environmental implications of biotechnology.
In this debate, risks involved in the use of biotechnological approaches have often been
emphasized (or even overemphasized) and the adequate guidelines for safety have been
suggested ad enforced by law. However, there have also been rapid developments in the
applications of biotechnology, which may help in controlling environment pollution, thus giving
a cleaner and sustainable environment in future. According to one estimate in USA, the US
market for environmental clean-up applications was expected to grow at an average rate of 17%,
while that for microbes and enzymes was expected to grow by only 7% every year.

Besides others, these applications for environment clean-up include biotreatment methods for
effluents and toxic wastes (this subject is described as bioremediation and is discussed in the
next). However, these bioremediation treatments, it is feared, could be problematic, where they
involve deliberate or accidental release of genetically modified microbes to the environment.

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READ THIS CAREFULLY. NEXT TIME YOU ARE ASKED TO PEE IN A CUP IT MAY
NOT BE FOR YOUR BENEFIT!!!!!!!!!

http://www.molecular-plant-biotechnology.info/transfection-methods-and-transgenic-
animals/transfection-methods-and-transgenic-animals.htm

Transfection Methods and Transgenic Animals - We discussed the methods used for cell,
tissue and organ culture in animals. Although, there is a variety of applications of animal cell and
tissue culture, one of its important application also involves transfer of foreign genes into our
livestock.

In organisms like bacteria and other microbes, or even in higher plants, the uptake of genes by
cells is often described by the term 'transformation'. However in animals this term has been
replaced by the term 'transfection', because the term 'transformation' in animal cell culture is used
to describe phenotypic alteration of cells.

The usage of the term 'transformation' for 'cell alteration' has been unfortunate and
discontinuation of its usage for this purpose is suggested.

Transfection or gene transfer in animals may be carried out at the cellular level to get transfected
cells, which may be used for a variety of purposes such as production of chemicals and
pharmaceutical drugs.

It may also be undertaken for basic studies involving study of structure 'and function of genes.
Although many mammalian cell lines have been regularly utilized for these purposes,
transfection has also been achieved successfully for the production of transgenic animals.

The improvement in livestock through transgenesis has already led to the following encouraging
results:

(i) increased milk production in cattle;

(ii) increased growth rate of livestock and fish

(iii) large scale production of valuable proteins in milk, urine and blood,
of livestock, enabling the use of transgenic animals as 'bioreactors' for
'molecular farming'.
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FOR THE DATA DOGS THAT FOLLOW ME ALL THE WAY DOWN THE RABBIT HOLE
HERE IS YOUR BONE:
CAULIFLOWER MOSIAC VIRUS IS ESSENTIAL TO THE EXPRESSION OF GMO
MANIPULATION IN ALL OTHER LIFE FORMS. START DIGGING:

http://www.molecular-plant-biotechnology.info/plant-viral-vectors/structure-and-properties-of-
CaMV.htm

Two major RNA transcripts are found inside infected cells. One covers the sequence of ORF VI.
Another and larger one contains the base sequence of the whole genome. (The RNA polymerase
goes all the way around the template strand and then continues for a short distance, repeating the
first transcribed segment.) It is likely that the larger RNA acts as a polygenic messenger RNA.

This suggests that any gene inserted into the genome will be transcribed.
Genes inserted in the correct direction may well be translated; however, it is possible that the
genes contain some special information for initiating translation besides the base triplet AUG.
This special information may be something like the Shine Delgarno sequence, even though this
sequence is not found in most eukaryotic mRNAs.

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http://www.molecular-plant-biotechnology.info/plant-viral-vectors/use-of-CaMV-as-vector.htm

Use of CaMV as Vector - From our knowledge of GaMV's structure and its infection process,
we can guess that this virus might help researchers to introduce genes into plants in a way that
would encourage their spread and expression. GaMV has several features in complete contrast to
those of Ti plasmid, some of which make it quite attractive as a vector.

One useful feature is that the naked DNA is, infective, being able to enter plant cells directly if
rubbed onto a leaf with a mild abrasive. Once inside the cells, the DNA is replicated and
encapsidated within virus particles, which then invade the rest of the plant.

Although the CaMV DNA does not become integrated into the chromosomal DNA and is
therefore not certain to be handed on to all cells during cell division, its spread throughout the
plant means that transformed plants can be effectively cloned by vegetative propagation.

But there are several problems associated with the use of CaMV as a gene cloning vector for
plants. Firstly, the genome is so tightly packed with coding regions that there is little room to
insert foreign DNA. Most deletions of any significant size destroy virus infectivity except for
small modifications in a specific region.

Inserts up to 0.4 kbp long are tolerated but those over 1.3 kbp destroy infectivity of the DNA.
Attempts have been made to sidestep this size limitation problem by using a helper virus system,
wherein a sub stantial proportion of the viral genome is deleted and replaced with foreign
DNA.The loss of function could be complemented by coinfection with a normal viral DNA, or
viral DNA deleted for a different function. However, the rescue of viral functions in all
experiments has occurred by recombination between the inactive viral genomes and only normal
infectious virus recovered.

For a helper virus system to be of any use, the recombinational rescue of altered genomes must
be suppressed, albeit the "retroviral like" mode of replication produces a high recombination
frequency and alteration of this would affect viral replication.

Secondly, the infection, once established, becomes systemic, spreading throughout the whole
plant. This lack of inheritance through the germ line might be advantageous in that the CaMV
DNA, and any inserted gene sequence, would be highly amplified in the host plant cells,
potentially permitting the expression of large quantities of the foreign gene product.How ever, it
appears that to propagate CaMV and to allow its movement through out the vasculature of the
plant, the DNA must be encapsidated; this would impose serious constraints on the size of
foreign DNA to be inserted into the viral genome.

Thirdly, CaMV DNA has multiple cleavage sites for most of the commonly used restriction
endonucleases. This would limit the usefulness of wild isolates of GaMV.

To date, the infectivity of the virus particle and its naked DNA, are the most useful assets in
terms of GaMV utility and development as a gene cloning vector for plants.

Not only does CaMV have a unique mode of replication, it also displays a peculiar translational
coupling of at least three genes that imposes some serious constraints on the use of the virus to
express foreign genes in plants.

Only small genes that do not exceed 300 base pairs in length and are devoid of introns are stably
maintained and expressed by CaMV. The only gene of the virus that may be replaced by a
foreign gene is the one encoding the aphid acquisition factor, a protein necessary for the spread
of the virus in nature.

Genes that have been successfully expressed after introduction into plant cells by CaMV are a
bacterial dihydrofolic reductase gene and a human interferon gene. In addition, the CaMV
genome contains two strong promoters of gene expression in plant cells that are often spliced
into other vectors, including the Ti plasmid, to drive the expression of foreign genes in plants.

One report of the insertion of a foreign gene into a plant describes the use of GaMV. In this
experiment a gene from E. coli was inserted into the CaMV genome in place of a gene that was
unnecessary for viral replication. GaMV infected turnip cells demonstrated the presence and
expression of this gene in several ways.The gene chosen for introduction was the dihydrofolic
reductase gene (DHFR) from E. coli strain 67. The enzyme coded by this gene is very resistant to
the antibiotic methotrexate, whereas most other DHFR enzymes are inhibited by this compound.

The investigators, Brisson and his colleagues from the Frederich Miescher Institute in
Switzerland(1984) removed most of open reading frame II with restriction enzymes and ligated
the DHFRXN gene in the same place.

Brisson took particular care to reduce the number of excess intergenic bases between the DHFR
gene and the adjacent CaMV genes (I and III), which proved to be important for reproduction of
the virus.

Infected leaves, ones that developed several days after viral DNA was rubbed on older leaves,
contained the E. coli DHFR. This was demon strated by a Western blot, showing that the cells
had an enzyme protein that reacted with antibody to the E. coli enzyme. Uninfected leaves
lacked the antibody reactive protein.

The plants infected with virus were also resistant to methotrexate. They could incorporate 32p
into DNA in the presence of this drug and survive several spray treatments. Control plants were
unable to synthesize DNA in the presence of the drug and died.

This experiment shows the feasibility of inserting and expressing a foreign gene, in fact a
prokaryotic gene, in a plant cell. As we might expect, the gene was present only in infected cells
and there was no evidence that it moved into a stable position in the plant genome.

In fact, when a larger piece of DNA containing the DHFR gene was inserted into the CaMV
DNA, it was unstable even in the viral genome and tended to disappear, being undetectable in the
progeny virus.

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THIS IS OF MAJOR SIGNIFICANCE BUT OF WHAT I DON'T KNOW. THE FACT THAT
mDNA IS BIGGER IN PLANTS HAS TO BE IMPORTANT. BEING A VEGETARIAN
CLEARS YOUR MIND.

http://www.molecular-plant-biotechnology.info/mitochondrial-genome/mitochondrial-
genome.htm

The plant mitochondrial genome has long been an enigma to molecular biologists. Even the
smallest is more than 200 kilobases in size, more than 10 times the size of animal mitochondrial
genomes (15-18 kb) and several times the size of mitochondrial genomes in fungi (18-78 kb) or
protists (15-47 kb). In contrast to the DNA of chloroplasts which is relatively conserved, the
DNA of mitochondria exhibits a wide variation in size and form.

Higher plant mtDNA can be circular or linear and varies from 200 kbp (in brassicas) up to
greater than 2500 kbp (in muskmelon).

The genome of plant mitochon dria is thus very large and may also be divided between one or
more DNA molecules.
The largest plant mitochondrial genome studied so far is half the size of the entire E. coli genome
(4,500 kb). The large size of these genomes presents several problems.

On the one hand, simply determining the physical structure of such large genomes is a major
challenge, especially since recombination events are known to produce several different
molecular configurations.

On the other hand, the large size per se and the dramatic variation of such genomes demands an
explanation.

We know of few genes in plant mitochondrial genomes which are not also present in the
mitochondria of yeast or animal cells.

And the few additional genes we do know about do not begin to account for the additional DNA
in even the smallest of plant mitochondrial genomes. Both animal and plant mitochondria encode
their own ribosomal and transfer RNAs.

The number of proteins encoded in plant mitochondrial DNA is probably not much higher than
the number encoded in mammalian mitochondria.

Experiments have shown that most of the protein synthesis in isolated maize mitochondria can be
accounted for by some 18-20 polypeptides.

Although there is always a possibility that more proteins might be synthesized in mitochondria in
vivo than in vitro, it seems likely that most of the mitochondrial genome is noncoding DNA.

http://www.molecular-plant-biotechnology.info/plant-viral-vectors/plant-viral-vectors.htm

Plant Viral Vectors- Viruses provide natural examples of genetic engineering, since viral
infection of a cell results in the addition of new genetic material which is expressed in the host.

In both microbial and mammalian systems, viruses have played important roles in vector
development, so it is not unexpected that plant viruses are considered as candidates for plant
gene vectors.

Additional genetic material incorporated in the genome of a plant virus might be replicated and
expressed in the plant cell along with the other viral genes. In fact, a simple but economically
significant example of plant genetic engineering using a viral vector already exists.One
phenotype that is usually conferred on a plant by viral infection is called cross protection a plant
infected by one virus usually cannot be superinfected by a second strain of a related virus.

This phenomenon has been exploited in tomato green houses, where

persistent tobacco mosaic virus (TMV) infections can be a major
problem.
Before the introduction of resistant genes in the late 1970s, it was
common practice to inoculate tomato seedlings with a symptomless
TMV strain produced by nitrous acid mutagenesis. The mild strain
provided cross protection against infection by the more severe isolates
endemic to green houses.

BET YOU DIDN'T KNOW YOUR HOTHOUSE TOMATOES WERE VACCINATED!!!!!!

To learn more about TMV and its implications in Hu-man health check out
www.cafepress.com/icd999

Since my teenage years I have warned people about tomatoes/Love Apples, the atropine
containing Witch's Fruit that acts on the nervous system to promote MK. They were introduced
to a folk population that knew Nightshade family plants were poisonous by a fellow who ate
them in the public square and did not drop dead. This story is so much like the introduction of
digital TV that it is astounding. WHY would there be a crusade to get the population to
CONSUME (eat a substance or watch TV) if there was not an agenda behind it? Witches are
SKILLED chemists andbotanists.

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http://www.molecular-plant-biotechnology.info/plant-viral-vectors/geminiviruses.htm

Geminiviruses - As with the Ti plasmid Agrobacterium and cauliflower mosaic virus, the
potential of geminiviruses as gene-cloning vectors for plants, stems from the work on several
plant diseases now recognized as being caused by these agents.

Both curly top virus (CTV) and maize streak disease virus (MSV) are geminiviruses
characterized on the basis of their unique virion morphology and possession of single stranded
DNA. Geminiviruses have a much wider host range than CaMV

http://www.molecular-plant-biotechnology.info/plant-viral-vectors/caulimoviruses.htm

Caulimoviruses - The caulimoviruses group consists of 6-19 viruses, each of which has a
relatively limited host range; the commonest. The best known member, cauliflower mosaic virus
(CaMV), infects many members of Cruciferae (cabbage, cauliflower, turnips, brussel sprouts,
rapeseed, Arabidopsis, etc.) and Datura stramonium.

Virulence of different CaMV strains varies from very mild (essentially latent) to lethal. The virus
is readily transmitted mechanically and by aphids; there is no evidence for seed
transmission.Caulimoviruses are similar in particle size, in vivo behavior and several are
serologically related.

They are confined to a few closely related plants in nature. There appears to be little, if any,
overlap between the host ranges of the individual viruses within the group, in spite of some of
the close serological affinities.

CaMV is experimentally transmissible to a few plants outside the Cruciferae family.

Caulimoviruses are widely distributed throughout the temperate regions of the world and are
responsible for a number of economically important diseases of cultivated crops.

The impetus for development of CaMV as a vector stems from study of the virus itself, as a
consequence of its pathogenic activities on susceptible plants. Symptoms of infection vary,
depending on the virus isolate, time of inoculation and condition of the plant, from mild vein
clearing to more severe leaf stunting.

The virus is transmitted by aphids in a nonpersistent or style borne manner. Successful transfer
of CaMV by aphids requires the presence of a transmission factor in infected cells. This factor is
not part of the virus particle but must be synthesized in response to infection since two non
transmissible isolates of CaMV have been identified.

However, one of the main attractions is that both the virus and the isolated DNA are infectious
and readily transmitted by abrasion of the leaves. In laboratory experiments virus particles (or
the DNA isolated from them) have infected a plant when rubbed on the surface of a leaf in the
presence of a small amount of abrasive compound.

Upon infection, the virus particles reproduce in the plant cells. Their DNA genomes serve as
templates for the synthesis of new DNA and for the transcription of messenger RNA, which
encodes for coat protein and other proteins that might be needed for the maturation and spread of
the virus.

In an infected plant virus particles move through the phloem into the expanding leaves, where
they disturb leaf development. These leaves have a mosaic or mottled appearance, with vein
clearing and dark green islands in the midst of more normally colored regions.

Differences in the expansion rates of different leaf regions lead to a crinkled and stunted leaf
appearance. How the virus produces changes in leaf morphology is itself an interesting problem
but since the same symptoms are caused by many other viruses, it is unlikely that unravelling the
specific characteristics of GaMV will help much in solving this problem.

In infected cells GaMV particles accumulate in cytoplasmic inclusion bodies. Early labeling
studies showed that radioactivity accumulates rapidly in these inclusion bodies, suggesting that
viral replication occurs there. How ever, recent evidence from several labs shows that replication
and transcription of CaMV probably occur in the nucleus.

Transcription of GaMV in isolated nuclei is 8-amanitin sensitive, suggesting that RNA

polymerase If transcribes the genome. Isolated nuclei from infected plants can incorporate a
radioactive label into viral DNA.

A covalently closed form of GaMV without gaps can be isolated from infected cells; probably
this molecule is packaged in nucleosomes. The host range and other biological properties of
CaMV suggest that it is not likely to become an economically important vector in future.
However, as a DNA virus whose genome is known to be packaged in nucleosomes and
transcribed by RNA polymerase II, it is presently more suited than, any other plant virus for
exploitation as an experimental tool, in the same way as SV40 and polyoma have been used in
mammalian systems. The knowledge gained from a GaMV model system should contribute to
the development of other types of plant vectors.

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BINARY WEAPONS DELIVERY SYSTEMS:

Dr. C and I just had a discussion on Helper Bacteriophages and Satellite Viruses in plants that
gave a Master Template for the Holy Virus weakening the mind so that it was susceptible to the
Evil Virus.

http://www.molecular-plant-biotechnology.info/plant-viral-vectors/structural-features-of-
geminiviruses.htm

Structural Features of Geminiviruses - The most surprising features of this virus group are the
small capsid size, 18-20 nm x 30 nm, their geminate (paired particles) morphology, which sets
them apart from all other classes of viruses, and the unexpected covalently closed circular
topography of the single stranded DNA which is in the molecular weight range of 7 x 105 to 9 X
105.

All geminiviruses recognized so far have a single major coat protein subunit in the range of 2.7-
3.4 x 104 daltons. The genome of the geminiviruses consists of either one or two circular, single
stranded DNA molecules.

The single stranded viral DNA, 2.6 to 3.0 kb long, is converted in the nucleus of plant cells into a
double stranded replicative form by an as yet unknown mechanismMany copies of the replicative
form of a geminivirus genome accumulate inside the nuclei of infected cells. There is 110
evidence to date for a reverse transcription step in geminivirus replication.

Bean golden mosaic virus (BGMV) DNA was found to be 2510 nucleotides long; if this was the
complete genome, it is less than half the length of any other known autonomously replicating
plant virus.

By comparing the single stranded DNA of the virus particle with the viral double stranded DNA
found in infected plants, it was found that the nucleotide sequence had a complexity twice that
expected given the physical size of the viral DNA.

This indicates that the BGMV DNA is heterogeneous and has a divided genome consisting of
two DNA molecules of approximately the same size, but differing in genetic content.

It would appear that geminiviruses consist of two populations of paired particles, differing only
in the nucleotide sequence of the DNA molecules they contain. Transmission of the virus in
nature occurs by leafhoppers or the tropical whitefly.
THEY HAVE BEEN PUTTING BACTERIAL PLASMIDS INTO VIRUSES THEN INTO
PLANTS. DO YOU REALLY THINK NONE OF THIS IS IN THE BIOSPHERE?
http://www.molecular-plant-biotechnology.info/plant-viral-vectors/use-of-geminiviruses-as-
cloning-vector.htm

Use of Geminiviruses as Cloning Vector - One advantage this group of viruses does have is
that they contain DNA which, although single stranded, appears to replicate via a double-
stranded intermediate, which would make in vivo manipulation in bacterial plasmids more
convenient.

The virus group is known to infect a wide range of crop plants, including monocots and dicots.
Attempts are underway to develop wheat dwarf geminiviruses as vectorsA potential
disadvantage may relate to the observation that in infected plants BGMV particles are limited to
phloem associated elements. Also these viruses are not readily transferred by mechanical means
from plant to plant; they are transmitted in nature by insects in a persistent fashion.

The small particle size may present packaging problems for modified DNA molecules.
Geminiviruses exhibit a much wider host range than do caulimoviruses. This makes the
geminiviruses more suitable for general application as vectors for introducing new genes into
plants.

Although little is known about the geminivirus genes and their functions, recent evidence
suggests that the gene encoding the viral capsid protein may be replaced by foreign genes
without interfering with the replication of the virus genome.The cereal geminivirus, wheat dwarf
virus, is under development as a vector for introducing genes into cereals. Although the single
chromosome of the wheat dwarf virus is only 2.7 kb long, it is nonetheless capable of
accommodating and replicating gene inserts up to about 3 kb in length. In this way the size of the
viral DNA may be more than doubled.

Three bacterial genes that have been inserted into the wheat dwarf virus genome have been
successfully replicated and expressed after transfer into cultured cells of the cereals Triticum
monococcum (one grained spelt) and Zea mays (maize). This illustrates the potential of
geminiviruses for serving as the basis of autonomously replicating expression units in plants.

http://www.molecular-plant-biotechnology.info/plant-viral-vectors/host-range-of-some-
caulimoviruses.htm

Host Range of Some Caulimoviruses -

Serological
Virus Host Range
Relatedness
Carnation etched ring Related to CaMV
Caryophyllaceae.
virus (CERV) and DaMV
Cauliflower mosaic Several Cruciferae and two species of Related to CERV
virus (CaMV) Solanaceae and DaMV
Dahlia mosaic virus Several Asteraceae, some Amaranthaceae, Related to CaMV
(DaMV) Chenopodiaceae and Solanaceae and DaMV
Unrelated to CaMV/
Mirabilis mosaic virus Mirabilis sp.
DaMV
Strawberry vein
Fragaria sp. ---
banding virus

http://www.molecular-plant-biotechnology.info/transgenic-plants/transgenic-plants.htm

Transgenic Plants - We discussed a variety of methods which can be used for transfer of foreign
genes to plant cells, tissues or organs. This transformation has been achieved at the level of
protoplasts or cells in many plant species although the ultimate objective should be the
production of transgenic plants following regeneration of whole plants from transformed
protoplasts/cells.

Not in all cases, the success in transformation could be combined with success in regeneration.
However now there are more than 50 plant species, where transgenic plants have been
successfully produced. Initially, the production of transgenic plants was restricted to
dicotyledons, but it has now been extended to several monocotyledons like wheat, maize, rice
and oats.

Progress in this exciting area of research for production of transgenic plants has been so
spectacular that by the turn of the century, we hope to be growing crops which have been tailored
to market specifications by the addition, subtraction or modification of genes.

Transgenes will also be important in increasing the efficiency of crop production systems.
For instance, transgenic plants resistant to herbicides, insects, viruses and a host of other stresses
have already been produced. Transgenic plants have also been produced, which are suitable for
food processing (e.g. bruise resistance and delayed ripening in tomato).

Another exciting example is the production of male sterile (due to barnase gene) and fertility
restorer (due to barstar gene) plants in Brassica napus, so that hybrid seed in future will be
conveniently produced without manual emasculation and controlled pollination as practiced in
maize.

This has also eliminated the need for a search of cytoplasmic male sterility (cms) and fertility
restoration system in crops, where hybrids are intended to be produced for higher yields.

Another major goal for production of transgenic plants, is their, use as bioreactors or factories for
production of speciality chemicals and pharmaceuticals. This area is described as molecular
farming or molecular pharming. The transgenic plants have also been produced for identification
of regulatory sequences for many genes, using gene constructs with overlapping deletions.