animals Flashcards
discuss 8 examples of transgenic animals
discuss issues with one of these examples
1)AquAdvantage salmon
a regulatory gene for growth hormone from pacific salmon was introduced along with a metalliothionine promoter from a pout
salmon grows all year around rather than just in spring and summer; the salmon is at market size in 1.5 years rather than 3. this is good because there is an increasing global demand for fish which cant be met by natural populations
30 years to commericialise; observations submitted before being reviewed by FDA, then public
concerns; fish are particular risky in terms of gene escape/transfer; fihs cant be rounded up easily.
fish may outcompete wild type fish and alter ecological composition
toxic products may effect consumers
people want food to be organic; little incentive for investment
to deal with this, AquBounty ensured fish are sterile and kept in land based farms so the risk of escape is minimised
2) enviropigs which can beter utilise phosphorus in animal feed due to engineering of phytase enzyme into salivary glands
70% of phosphorus is in phytic acid form which is indigestible to pigs and is excreted which causes environental problesm
this is a great solution because farmers dont have to add microbes capable of breaking down phytase or phosphate to feed.
also the phytase in manure was reduced by 60% which is good for the environment
project gained FDA approval but was shut down due to lack of commercial interest (people dont want GM foods)
3) Oxitec mosquito is a male mosquito which is engineered to carry a self limiting gene (needs an antidote to survive) when these males mate with females, the gene is inherited by offspring which cant survive into adulthood
this can be used to control the spread of malaria and zika
trials have shown a 90% reduction in mosquito population
there is always concern with releasing GM animals into the wild but the mosquito does have a florescent red marker gene its populations can be tracked
benefits would include less pesticide needed, less human deaths, less economic costs
4) cows which prodcue more milk or milk ith less lactose and cholesterol
5) sheep with more wool; less animals needed to meet demand
6) animals with more meat
7) in 1982 the first transgenic animal was created. the promotor for growth hormone was changed to a zinc sensitive hormone. when the mouse was fed a high zinc diet the mouse grew very large.
8) pharming; using animals to produce pharmacuticals.
in 2006 goats producing anti-thrombin (prevents blood clots) were approved; first ever GM animal product approved
microbes can also be used to produce pharmacuticals but animals are better if larger amounts are needed or if the protein is complex and glycoslated
disucss the timeline of approving a GM animal with reference to a case study; 5 marks
1985; first transgenic fish produced; expressed human growth hormone. unsuccessful as the genes from fish and humans were too different to eachother
1990’s (5 years); salmon produced with metallothionine promotors; very successful
1996 (11 years); findings on observations submitted
2010 (25 years years later); FDA said it was unlikely to cause environmental effects
2013 (28 years later); public comments taken and salmon reviewed again
2016 (31 years later); canada approved sale of salmon
why is animal transformation considered more difficult than plant transformation
plants are very flexible
animal modification must be carried out on germ cells (female eggs are already developed and present at birth) so modification must be done on early embryos
discuss which animal is used for GM trials and why
mice
easy and cheap to keep because theyre small
reproduce very quickly so inheritability can be investigated
mammals so results can be generalised to humans. all mamals have similar genes for early development
human disease genes can be expressed into mice to study human diseases
discuss the two methods of creating a transgenic mouse.
reference advantages and disadvantages of each method
pronuclear injection
1) a gene construct is engineered which includes a promotor region, an intron between promotor and exon which allows for Tf binding, polyadenylation sequence so that the mRNA has a polyA tail
2) the gene construct is is engineered into bacteria so that lots of copies of the gene are produced. the plasmid is removed from the bacteria and cleaved so that its linear
3) a newly fertilised egg is collected (before the maternal and paternal nuclei have fused
4) a suction pippette is used to hold to the egg in place while a micro pipette is used to insert multiple copies of the transgene into the male pronuclei. suction pipette is needed because mammal eggs are very small and hard to handle
5) egg is inserted into a pseudomother and a sterilised male is used to induce bodily responses needed for pregnancy
6) pieces of mice tail are taken and PCR is done to check the trangene has been incorporated
7) so who do contain the transgene are bred with wild type mice to check if the offspring have the gene.
drawbacks;
1) genes can only be inserted, not modified
2) random site of insertion; position effects and gene disruption is possible
3) multiple copies may be inserted rather than one
4) the trangene is not always consistently inherited, hence the followign method is used.
homologous recombination
1) a gene construct is created including GOI and marker gene
2) blastocyst cells are harvested and cultured to form a mass of embryonic stem cells which are totipotent
3) methods such as electroporation (electric current opens voltage gated channels in plasma membrane) and lipofection (attach liposomes areound DNA and then fuse with plasma membrane) are used to allow the trangene to enter the nucleus
4) the trangene inserts itself into a precise location due to pieces of DNA on either end of the GOI known as the targetting vector which are homologous to pieces of DNA
5) crossing over occurs; the complimentory strand synthesises a complimentory strand
6) the embryonic stem cells are cultured on a selective medium which only allows transformants to grow (positive selection; only those with antibiotic resistance gene grow). to ensure the targetting vector isint inserted it often contains a toxin gene (negative selection)
7) ES cells are placed inside another blastocyst, which in placed in a pseudomother
8) the offspring is a chimera (featueres from both wild type and from transformed blstocyst). chimera bred with wild type to see which chimeras have germ cells which developed from the ES cells (easy to tell because ES cells come from a mice of one colour, and blastocyt comes from a mouse of another colour). those with the colour relating to the ES cells are selected for
positives: only one copy of the gene is inserted, allows for modification of preexisting genes not just insertion, potential for knock in insertion which doesn’t effect any prior genes. more consistantly inherited by offspring
negatives; efficiency is very low
what does knock in and knock out mean
what is an application of knock out
knock in; insertion of a transgene into an organism without effecting any prior genes
knockout; insertion of a transgene into an organism in a place resulting in inactivation of a pre existing gene
knockout can be used to study the function of a gene
