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Genetic Engineering

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Biology

Genetic engineering is a more focused form of selective breeding that humans have been performing for thousands of years. While selective breeding focuses on breeding two organisms with desirable phenotypes together, genetic engineering focuses on directly manipulating an organism's DNA to change its phenotype. Selective breeding changes the DNA indirectly whereas genetic engineering directly changes the DNA of an organism.

What is genetic engineering?

Genetic engineering, as previously mentioned, builds upon the selective breeding that humans have been using for thousands of years to reinforce desirable phenotypical characteristics within organisms. By breeding organisms with the desired characteristic together past humans hoped to reinforce this phenotype or create new ones.

Some classic examples of selective breeding include the numerous dog breeds and modern grains we have today, which evolved from wild wolves and grasses respectively.

To learn more about this first form of indirect genetic engineering read our article Selective Breeding!

Instead of breeding organisms together and hoping the desired outcome occurs, genetic engineering takes influencing phenotypes to another level by making the desired changes directly to the DNA (genotypes). These changes are performed through a variety of techniques, mostly involving the use of recombinant DNA technology. The organisms that result from this genetic manipulation are known as genetically modified organisms.

Recombinant DNA technology is a very important research tool in biology. It consists of several laboratory techniques that allow scientists to manipulate, isolate and combine different DNA fragments of interest to create genes with new functions and produce desirable proteins. This so-called recombinant DNA is then usually transferred into a bacterial or yeast cell where its copied and expressed.

The Genetic Engineering Process

The process of genetically modifying an organism or a cell involves several key steps, each of which may be achieved through a variety of means, however, whatever the mechanism used the end goal is the same.

Selection Of A Target Gene

Genetic engineering begins with the identification of the genes the scientist wishes to insert into the genetically modified organism. Which genes are required depends on what we want the organism to do. Where the trait is controlled by a single gene only one gene needs to be moved. This is the simplest way to insert a new trait into a genetically modified organism. An example of this is the insertion of the insulin gene into Escherichia coli for the production of human insulin.

Many phenotypical traits are however polygenic or the result of the action of many genes. These include a large number of metabolic pathways, amongst other traits. These are more complicated to use in genetic engineering as each gene responsible for the traits, or at least enough of them to produce the desired part of the trait, must be identified.

Genes may be identified using either forward or reverse genetics. Forward genetics identifies the desired phenotype first, then finds the gene or genes responsible for it. By inserting DNA sequences into the genome at known locations and looking for changes in the phenotype, allowing the phenotype to be linked to that gene. Reverse genetics functions the opposite way. Thanks to modern genetics techniques, large amounts of sequence data are often available early on in the research process. This allows for the identification of genes early, which may then later be linked to phenotypes through introducing mutations or a variety of other means.

Manipulation Of The Desired Gene

Once the desired gene is identified it must be extracted from the organism ready to be inserted into another. The gene must then be manipulated in order to ready it for insertion into the genetically modified organism that is to be created. This involves the modification of the gene to allow it to be expressed by the organism it is to be inserted into.

Gene Extraction

The gene must be extracted from the cell in order to be further manipulated. Although In vivo modification of DNA is possible, it's also harder. The cell is first ruptured in order to release the contents into the solution. This is done either mechanically, such as by grinding or even freezing or chemically breaking the plasma membrane. The DNA is then purified, with the necessary steps depending on the lysis method used, ready for downstream uses.

Gene Isolation

In order to insert the gene into another organism, it must be separated from the DNA that makes up the rest of the source organism's genome. How this is performed is dependent on whether the sequence of the gene is known or not. The most widespread technique uses restriction enzymes to isolate the gene and other proteins to insert it into a vector like a bacterial plasmid or a virus. This process is known as molecular cloning!

Read our article Artificial Cloning to learn more about how molecular cloning happens!

Restriction enzymes are very important tools for genetic engineering and recombinant DNA technology. These bacterial proteins cut DNA from all organisms at a sequence-specific site, leaving DNA fragments with known sequence ends. This allows scientists to manipulate and insert these DNA fragments easily. There are over three thousand of these enzymes each recognising and cutting a specific DNA site!

Vectors are molecular biology tools and DNA molecules (usually bacterial or viral) used to facilitate the transfer of other DNA into a host cell. The vectors are the vehicles in molecular cloning! They also usually help with expressing and replicating the exogenous DNA in the host.

Gene Insertion

In order for the genetically modified organism to express the desired gene, the now modified gene must be either inserted into its genome or as extrachromosomal DNA. The transmission of the modified gene to offspring can be controlled by controlling where the gene is inserted. Insertion into somatic cells means that the gene cannot be passed on, but insertion into germline cells allows it to be passed down.

Before the gene can be inserted into its expression system it must be modified to allow efficient transcription and translation of the gene. Generally, some form of marker is added to allow for the identification of successfully modified organisms, such as antibiotic resistance genes or fluorescent proteins. A promoter region that encourages the transcription of the gene is also usually added in order to increase the expression of the desired phenotype. A terminator region is used to instruct the expression system to terminate transcription.

There are several techniques used to physically force the insertion of external DNA into a host cell. Each technique has different specificities. The most famous techniques include:

  • Transformation - Insertion of DNA in bacteria using recombinant DNA plasmids.
  • Transduction - Recombinant viral vectors are used to insert genes into mammalian cells.
  • Transfection - Insertion of small molecules like DNA or RNA into eukaryotic cells via non-viral means.

Biolistic transformation is an unusual type of transformation where nucleic acids are shot into organisms in high-speed particle bombardment! Heavy metal particles are coated in the gene, and then literally fired into the cell using a gene gun, carrying the gene with them.

There are several different types of transfection methodologies. It may be done via physical means, which force the genetic material into the cell, or by introducing pores (electroporation) into the membrane through which DNA may enter. Chemicals may also be used to trigger the entry of the genetic material into the cell for example in lipofection.

Targeting Of Gene Insertion

Most techniques used to genetically modify organisms insert the desired gene at random places within the genetic material of a cell. While this may work fine and have no negative effects it may also result in disruption of genes within the organism and consequently their function which can affect the cell. More recently programmable enzymes have been developed to allow precise genetic modifications. These enzymes known as nucleases create double-strand breaks within the DNA at specific points, allowing DNA repair mechanisms to be hijacked to insert DNA at that site. These nuclease systems include TALENs, zinc finger nucleases (ZFNs) but especially CRISPR/Cas which is revolutionizing genetic engineering and the world!

CRISPR/Cas stands for Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR associated protein (Cas). CRISPR/Cas is an RNA/protein complex that was discovered as part of the bacterial immune response against viral infection. Its current fame stems from the fact that it's the ultimate molecular scissors! CRISPR/Cas is essentially the best gene-editing tool available, allowing us to precisely introduce base-specific changes in DNA.

Uses And Examples Of Genetic Engineering

Genetic engineering has many uses both in and out of the laboratory in many fields such as agriculture, research, industry, medicine, novelty items, and conservation.

Medicine

In medicine, genetic engineering has been used for a variety of purposes. One of the main examples is the production of drugs such as the bacterial production of human insulin, which was the first example of the use of bacteria to mass-produce human biological molecules. Currently, these techniques are used to create many things, including monoclonal antibodies, hormones, vaccines, coagulation factors and other proteins.

By modifying animals to have certain genetic diseases, the underlying causes of these diseases can be studied and cures tested. Modifying animal genetics may also remedy the shortage of organs for transplantation by increasing the compatibility of animal organs during xenotransplantation. While genetic engineering may be used to allow cures to be tested, it may also be used as a curative in itself. Gene therapy is used to genetically modify humans to correct genetic diseases such as sickle cell anaemia, Duchenne's Muscular Dystrophy (DMD) and leukaemia.

Gene therapies are genetic engineering techniques that use genes to directly treat or cure an inherited or acquired genetic disorder. With the advent of the CRISPR/Cas, gene-editing based gene therapy is becoming increasingly popular and successful at directly correcting base-specific disease-causing mutations.

Research

Genetically engineered organisms may be used in research. By inserting genes into bacteria, a near-infinite supply of them may be easily and cheaply provided. Other avenues of research centre around modifying the function of genes within an organism to study the role and functions of the gene. The modification may involve alterations to the function of the gene, tracking of the protein the gene encodes or analysis of the gene expression.

Novelty Products

Genetic engineering has also been used to produce several novelty products, which usually stem from rebranded organisms that were created as part of research projects, such as GloFish, which are fish genetically engineered to express fluorescent proteins in a range of colours. They were initially developed in a lab at the National University of Singapore as part of a research project aiming to create a fish that could detect environmental pollutants and fluoresce. Other examples of novelty genetically modified organisms include bacteria modified to be able to create art, glowing plants and new coloured flowers.

Industry

Genetically modified organisms are used in industry to create large amounts of desired proteins, in a similar manner to how they are used to create many medications. They have also been used in waste management, including feeding on oil, plastics, extracting heavy metals and other contaminants from land and many other uses. They have also been used to create materials including carbon nanotubes and lithium batteries. They are also used in the production of food and as sensors for numerous environmental factors.

Agriculture

The use of genetically modified organisms in agriculture is a very controversial topic, however, it provides many opportunities to solve large issues facing society. Plants have been modified to become more resistant to pathogens, be more nutritious, have larger yields, produce different compounds, grow faster or better resist environmental conditions.

Conservation

Genetic engineering has been used in conservation to allow plants to resist pathogens, create viruses that can control invasive species or vaccinate native organisms. It could also potentially be used to help native organisms adapt to climate change and other threats.

Benefits And Risks Of Genetic Engineering

There are a number of advantages and disadvantages of genetic engineering, which are surmised below.

Advantages Of Genetic Engineering

  • It allows organisms to be modified to do exactly what scientists need.
  • It solves reliance on other potentially more destructive technologies, like pesticides.
  • It allows the production of substances otherwise difficult, expensive or impossible to manufacture in large volumes.

Disadvantages Of Genetic Engineering

  • The genetic modification may "escape" and enter other organisms.
  • The modification of organisms may have unintended consequences for the organism.
  • It raises several ethical and moral issues, such as can you patent life, and should we "play god".

Genetic Engineering - Key takeaways

  • Genetic engineering allows for direct changes to be made to the DNA.
  • This allows the enhancement of already occurring phenotypes or the addition of new ones to an organism.
  • The gene or genes responsible for the phenotype must first be identified then extracted and modified, before being inserted into the desired organism.
  • Genetic engineering may be used for medicine, research, novelty, industry, agriculture, conservation and many other areas.
  • genetic engineering is a versatile technique, however, it could have many unintended side effects.

Genetic Engineering

Genetic engineering, or genetic modification (GM) is when the genes of an organism are modified in order to produce an improved version of itself.

Scientists are able to identify specific genes that are desirable characteristics to have in organisms. When scientists find a gene of interest, they remove it from the cell using enzymes and directly insert this gene into the genetic material of another organism. 

Examples of genetic engineering include inserting human insulin genes into bacteria DNA, to produce a vast amount of insulin for human health, as well as modifying rice to contain beta-carotene, which is important for human nutrition.

Genetic engineering may be split into three types depending on either the method of genetic changes used, in which case there is conventional breeding, transgenic and cisgenic genetic engineering. They may also be subdivided according to the level of application, which gives you analytical genetic engineering, applied genetic engineering and chemical genetic engineering. 

Genetic engineering was first performed thousands of years ago, with humans selectively breeding plants and animals with desirable characteristics in order to enhance these. Direct manipulation of DNA for the purposes of changing an organism first began in the 1970's when Paul Bird combined the SV40 viral DNA with bacteriophage DNA. 

Final Genetic Engineering Quiz

Question

What is genetic engineering?

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Answer

Genetic engineering, or genetic modification (GM) is when the genes of an organism are modified in order to produce an improved version of itself.

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What is the role of insulin in human health?

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Insulin is a hormone which regulates blood glucose homeostasis.

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What is meant by the term ‘vector’?


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A vector is the method by which genetic material is transferred from the donor to the recipient. Examples of common vectors include bacterial plasmid, or a virus.

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How can GM crops with increased resistance to pests and disease outbreaks help to prevent environmental contamination or human health issues?


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Less herbicides and pesticides are used because GM crops are more resistant to pests and disease. Pesticides and herbicides cause environmental contamination, and can be harmful to human health.

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How can toxins produced by GM crops affect the ecosystem?


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GM crops may produce toxins that are harmful to pollinators (e.g. bees). This will affect pollination in the local environment, which could have wider consequences in the ecosystem.

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Genetic engineering makes what kind of changes to an organism's DNA?

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Direct

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Selective breeding changes DNA how?

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Indirectly

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Selective breeding breeds organisms with what together?

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Desirable phenotypical characteristics. 

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Genetic engineering uses what type of DNA technology?

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Recombinant

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How many main steps are there in the creation of a genetically modified organism?

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Three steps - gene identification, extraction and insertion.

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What type of enzymes are used in targeted gene insertion methods such as CRISPR/Cas?

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Nucleases

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What type of breaks do nucleases create in DNA?

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Double Strand

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What types of genetics are used to identify genes useful in genetic engineering?

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Forward

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Does forward genetics look at genes or phenotype first?

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Phenotype

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Does reverse genetics look at gene or phenotype first?

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Gene

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What is a classic example of genetic engineering?

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The production of human insulin in bacteria.

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What enzymes are used to extract the desired gene or genes from the genome?

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Restriction enzymes

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What technique other than restriction digestion can be used to separate the desired gene or genes from the genome?

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Polymerase Chain Reaction

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A promoter does what in a gene?

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Increases transcription of the gene.

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A termination region does what in a gene?

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Ceases transcription. 

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Why are markers added to genes?

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To identify successfully modified cells. 

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Transfection refers to the modification of what kind of cells?

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Animal

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Transduction uses what pathogen to insert genes?

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Viruses

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What is the difference between natural and artificial cloning?

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Artificial cloning uses lab-based manipulation to form a clone, whereas natural cloning is the formation of clones via natural means, often as a part of asexual reproduction. 

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What are methods used in whole organism cloning? 

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Somatic Cell Nuclear Transfer 

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Which of these mechanisms may result in the creation of a natural clone?

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Binary Fission

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Artificial cloning may be used for what purposes?

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Artificial cloning may be used for reproductive and therapeutic purposes.

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Reproductive cloning is done for what purpose?

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Creating more copies of an organism with desirable characteristics. 

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What form of twins are clones?

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Monozygotic

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What type of reproduction is used by animals to asexually reproduce? 

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Parthenogenesis, more specifically apomixis. 

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Binary fission is mainly used in what organisms?

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Prokaryotes

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What is gene cloning?

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Gene cloning is the cloning of genetic material. 

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What methods are used in gene cloning?

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PCR or replication within a host organism. 

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What is a key trait of cell lines?

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Immortality

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Cells may be immortalised using what methods? 

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Sourcing the cells from tumours, or the introduction of immortalising genes. 

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Plant tissue culture is also known as what?

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Micropropogation. 

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Somatic Cell Nuclear Transfer transfers the somatic nucleus into what type of cell?

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An oocyte or egg cell. 

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The first organism to be cloned from an adult organism was?

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Dolly the sheep. 

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Embryo splitting creates twins by?

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Manually splitting the blastocysts into two bundles of cells, which replicates how monozygotic twins occur in nature. 

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What sectors use genetic engineering?

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Medicine, research, agriculture, conservation, industry and novelty item production. 

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What are recombinant proteins?

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Proteins encoded by recombinant DNA. 

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Why are cell-based protein expression systems used instead of cell-free systems?

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Cell-based systems are much cheaper and more efficient than cell-free protein expression systems. 

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What is the downside of using bacteria to express complex proteins?

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They often are unable to perform necessary post-translational modifications and also often lack the necessary tRNAs to use the codons found in eukaryotic genes. 

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What was the first widely produced recombinant protein?

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Insulin

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Genetic engineering is used to do what in hybridomas?

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Modify the antibodies produced by the hybridoma.

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Genetic engineering allows scientists to model diseases. Why is this done?

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To bypass the limited population size of humans with certain diseases. 

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The main issues preventing widespread xenotransplantation are?

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Immunological challenges

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Gene drives ensure modified genes are passed on to what percentage of offspring and at what zygosity?

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100%, homozygous.

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Gain of function experiments result in what protein activity?

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Increased

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Loss of function experiments result in what protein activity?

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Decreased

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