Transgenic Animals and Plants
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Transcript of Transgenic Animals and Plants
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Transgenic Products of rDNA technology are produced by
genetic modification in which DNA coding for therequired product is introduced, usually by meansof a plasmid or a viral vector, into a suitable
microorganism or cell line or plant cell or animalcell, in which that DNA is expressed andtranslated into protein.
The desired product is then recovered byextraction and purification.
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Transgenic plants as factoriesfor biopharmaceuticals
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Molecular farming is the use ofcrops for the large-scale production
of valuable recombinant proteins.
Molecular farming in plants has arelatively short history, and the
commercial adoption of thistechnology occurred very recently.
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Glycosylation of proteins is a highly complex post-translational modification process taking place in
the endoplasmic reticulum and Golgi apparatusand involving more than a hundred differentproteins (and genes).
This glycosylation machinery is absent in E. coli,present but different from mammalian cells in the
yeast Saccharomyces cerevisiae, but highly
conserved among mammalian cells (e.g., human,Chinese hamster).
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Transgenic plants can also produce a variety ofproteins used in diagnostics for detecting humandiseases and therapeutics for curing human andanimal diseases in large-scale with low cost.
,proteins, peptide hormones and cytokinins arebeing produced in transgenic plants and theirparts such as tobacco (in leaves), potato (in
tubers), sugarcane (in stems) and maize (in seedendosperm).
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Plants are amazing and cheap chemical factoriesthat need only water, minerals, sun light andcarbon dioxide to produce thousands ofsophisticated chemical molecules with different
structures.
Given the ri ht enes lants can serve as
bioreactors to modified or new compounds suchas amino acids, proteins, vitamins, plastics,pharmaceuticals (peptides and proteins), drugs,
and enzymes for food industry and so on
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Producing therapeutic proteins in plants hasmany economic and qualitative benefits,
including reduced health risks from pathogencontamination,
comparatively high yields, and in seeds or otherstorage organs
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The cultivation, harvesting,storage,and processing of transgenic crops
would also use an existinginfrastructure and require relatively
e cap a nves men ma ng ecommercial production ofbiopharmaceuticals an exciting
prospect.
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Plants are potentially a cheapsource of recombinant products.
Estimated that the cost of
producing recombinant proteins inplants could be 10- to 50-fold lowerthan producing the same protein byEscherichia coli fermentation.
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The cultivation, harvesting, storage, andprocessing of transgenic crops would also use anexisting infrastructure and require relatively little
capital investment making the commercialproduction of biopharmaceuticals an excitingprospect.
In addition, purification may not always benecessary, for example, in the case of edible
vaccines. Plant-derived products, whetherpurified or not, are less likely to be contaminated
with human pathogenic microorganisms thanthose derived from animal cells, because plantsdont act as hosts for human infectious agents
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because they are easily transformed and provide a
cheap source of protein.
Several biotechnology companies are now actively
developing, field testing, and patenting plantexpression systems, while clinical trials are
proceeding on the first biopharmaceuticals derived
from them.
One transgenic plant-derived biopharmaceutical,
hirudin, is now being commercially produced inCanada for the first time.
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Techniques Used for Generating
Transgenic Plants
As with bacteria, the ability to geneticallymodify plants depends on obtaininggenetically identical populations and readily
manipulating DNA. How do you clone aplant?
It is also possible to grow plants in culture
from small explants.
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Another method is to culture plants fromtotipotent cells found in plant meristems.
These plant cells can divide and differentiateinto the various types of specialized cells. In atest tube, plant cells will divide and form an
.
When hormones in the culture medium areadjusted, the callus will sprout shoots androots and eventually develop into a plantletthat can be transplanted to soil.
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To clone a plant perhaps a plant withnew genes the growing callus issimply subdivided. Thousands of
genetically identical plants can begenerated in this way.
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Transformation transformed transgenic plants are produced using
Agrobacteriummediated transformation,
particle bombardment, or
other standard transformation techniques.
expression system, but other plants have been used,including
Nicotiana bethamiana, Arabidopsis thaliana,
tomato, banana, turnip, black-eyed bean, oilseedrape, Ethiopian mustard, potato, rice, wheat, andmaize.
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The second strategy is to infect nontransgenicplants with recombinant viruses that expresstransgenes during their replication in thehost.
used are
tobacco with tobacco mosaic virus (TMV) or
cowpeas with cowpea mosaic virus (CPMV)
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Expression of pharmaceutical proteinsin transgenic plants is mostly byconstitutive CaMV 35S promoters.
Promoter
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Seed expression has been achieved by placingtransgenes downstream of the glutelin (Gt3)signal peptide sequence and transcribing from
Gt3 promoters2.
Transcription of immunoglobulin transgenes intobacco has also been controlled b the seed-
specific, developmentally regulated, legumin B4promoter11.
As immunoglobulins in the cytoplasm often
interfere with cell function, it is desirable to havethem secreted into the apoplasm, or trafficked
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Trafficked to the endoplasmic reticulum,where they generally accumulate to a muchhigher proportion of TSP (15%)40. Traffickingcan e rec e y e egum n 4 s gnasequence
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Full-scale commercial production willprobably involve grain and oilseedcrops, such as maize, rice, wheat,soybeans, and oilseed rape.
Proteins stored in seeds become
esiccate , an cou remain intact orlong periods, making seeds a convenientmethod of storing, distributing, and
administering biopharmaceuticals suchas vaccines.
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Genetic Modification of Animals Dolly thelamb stole the headlines as the first example oflivestock cloned from DNA of an adult animal.
But the real breakthrough came with Polly, thefirst transgenic lamb. Born the year after Dolly,
P ll w iv n h m n n th t n
blood-clotting factor IX, the protein missing inpeople with one form of hemophilia.
Harvesting such proteins from transgenic
livestock is one goal of this research.
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The road to Polly and subsequenttransgenic animals began with researchusing genetically altered mice. Along the
way, technologies for cloning animals,modifying DNA, and targetingexpression of proteins to specific tissues
were developed.
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The production of pharmaceutical humanproteins in transgenic animals is still a minorbusiness, in which only a few companies are
.
has yet reached the market but the stewards ofthis technology hope to achieve this within thenext couple of years. Currently, most research is
directed towards products for industrializedcountries
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Transgenic animal
A transgenic animal is by definition an animalwhose genetic composition has been altered toinclude selected genes from other animals or
breeding. In other words, an animal altered by the
introduction of recombinant DNA through
human intervention.
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The first transgenic animal was a mouse, createdin 1981, carrying a gene which made the animalsusceptible to cancer.
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created in 1985.
Biotechnology firms and research institutesinvolved in pharmaceutical development and
production use transgenic animals for threedifferent ends.
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As apharmaceutical production unit. Viatransgenic technology such as micro-injection, genes that code for the productiono a uman pro e n are nser e n o egenome of an animal, which in turn producesthe human protein.
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Pharming
This new branch of transgenic animalproduction which has emerged in recent yearshas a new name:pharming. Pharming is thepro uc on o p armaceu ca umanproteins in transgenic farm animals.
In most cases of pharming, changing thecomposition of milk is the main strategy.
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Mammary glands of cows produce largevolumes of milk, a protein-rich solution whichcan be collected non-invasively. In October1997, a ure o ec no ogy repor e on eachievement of producing a biologically activehuman protein in the milk of a transgenic pig.This demonstrated the feasibility of
producing large and complex proteins in thisway.
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However, it remains difficult to generate animalswhich produce these medical proteins of aconsistent quality and in sufficient quantities. If a
successfully engineered transgenic animal can becloned, and then bred successfully, it will bepossible to create herds of these animals for
quality.Research also extends to areas other than milk.TheAgricultural Research Service of the United
States Department of Agriculture (USDA) hasdeveloped mice which produce stable amounts ofhuman growth hormone in their urine.
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Disadvantage
The researchers see some clear disadvantages of milkbased pharmaceutical protein production: firstly,lactation occurs only in females and is non-
,
months before lactation starts. Finally, milk is acomplex substance usually containing 3 to 6 per centtotal protein and therefore needs extensive
purification to obtain the pharmaceutical protein.Purification of urine seems to be easier, according tothe USDA research.
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Microinjection
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Why transgenic livestock The use of transgenic livestock in protein
production aims at overcoming several majorbarriers presented by cell-based systems.
o en a y, s approac cou prov e argequantities of complex proteins in a cost-effective way. Compared to the facilities andchemicals required for cell culture
production, capital investment required foranimal production facilities is relatively low.
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The British companyPPL Therapeutics (PPL)estimates that the basic costs for productsdeveloped by transgenic animals are four to ve mes ower an ce cu ure pro uc on.
However, this does not take into accountdevelopment costs. To date, the use oftransgenic animals is developing fast for a
range of pharmaceutical products.
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Drug Disease/Target Animal Company
alpha-lactalbumin anti-infection cow PPL
alpha1 anti trypsin(AAT)
deficiency leads toemphysema
sheep PPL
CFTR cystic fibrosis sheep, mouse PPL
R&D of medicine production by
transgenic animals
uman pro e n rom os s p g, s eep
tissueplasminogenactivator (tPA)
thrombosis mouse, goat PPL
human calcitonin osteoporosis rabbit PPL
factor VIII hemophilia pigsheep
PharmingPPL
factor IX hemophilia pig, cowsheep
PharmingPPL
Drug Disease/Target Animal Company
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g / g p y
fibrinogen wound healing cowsheep
PharmingPPL
alpha-glucosidase
Pompe disease rabbit Pharming
collagen I
collagen II
tissue repairrheumatoid
arthritis
cow
cow
Pharming
lactoferrinGI tractinfection,
cow Pharming
arthritisantithrombin 3(ATIII)
thrombosis goat GTC
glutamic aciddecarboxylase
type 1 diabetes mouse, goat GTC
human serumalbumin (HSA)
maintains bloodvolume
mouse, cow GTC
msp-1 malaria mouse GTC
Pro542 HIV mouse, goat GTC