To examine the questions surrounding gene drive research, this report relies heavily on an extended, iterative exploration of a set of plausible case studies. The case studies are first described in a preliminary fashion in this chapter. Other chapters build on these case studies with deeper discussion of issues pertinent to value-based concerns, scientific techniques to mitigate harms, risk assessment, public engagement, and governance.
The case studies offer practical scenarios on which to base the report’s analysis and recommendations and to provide a sound foundation for the further discussions that will necessarily follow this report as gene drive research advances. Given those two goals, the committee used the following three criteria to select case studies:
- Plausibility: Selection of organisms suitable for the development of a gene drive.
- Likelihood: Selection of areas for gene drive research or applications that are expected to be pursued in the near term.
- Diversity: Selections are intended to reflect a range of plausible target organisms, applications, mechanisms of action, and locations (in terms of where gene drive research is carried out and where organisms could potentially be released).
It is particularly important to understand what is meant by plausibility. Many organisms and traits are not suitable for gene drive research. The two most basic requirements for a target organism of gene drive work are that it reproduces sexually and that it reproduces rapidly (see Box 3-1). For this reason, many insects and rodents are good candidates for gene drive research. Organisms such as viruses, many plants, and most bacteria, which use other means to reproduce, are not good targets for gene drive research (see Box 3-2 for additional considerations for plants). Humans, elephants, and trees are also not good targets for gene drive research because they have long generation times; any modification introduced into such a population could require decades or centuries to become established. However, a gene drive could work in an organism that has alternating sexual and asexual phases of reproduction, as in Plasmodium falciparum, the parasite that causes malaria (de Koning-Ward et al., 2015), even though its population structure may render spread of the gene drive difficult.
In addition, some traits may simply be too complex to alter because they are governed by many genes, their expression is shaped by the external environment, or they are modified by internal or development cues (e.g., epigenetics) that are not yet fully elucidated. For example, flowering time in maize is determined by the cumulative effects of many genes (Buckler et al., 2009).
In some cases, many applications of gene drive research may not be necessary, because efficient non-gene drive approaches are able to generate the desired outcome.
Given these and other technical and regulatory challenges (discussed in detail in the other chapters), predictions about how gene drives might be used need to be treated critically. The committee developed case studies to illustrate the issues highlighted in Table 3-1.
Establish gene drives in Aedes aegypti and Aedes albopictus mosquitos to control the spread of dengue throughout the world.
Dengue, a debilitating viral infection, is one of the leading causes of sickness and death in subtropical and tropical countries around the world. Adults and children who contract dengue often experience a flu-like illness. Severe dengue, also called Dengue Hemorrhagic Fever, causes bleeding, persistent vomiting, breathing difficulties, and other complications that may lead to death. Severe dengue disproportionately affects children.
Dengue is caused by infection with any of five serotypes of dengue virus (which is a flavivirus). The virus is transmitted to humans via the bite of female1Aedes aegypti, the primary vector (carrier) in urban areas, or Aedes albopictus, the primary vector in rural areas. In April 2016, the World Health Organization endorsed the use of the first-ever dengue vaccine, Dengvaxia (CYD-TDF) by Sanofi Pasteur, in countries where dengue is endemic.2 Research is ongoing for other vaccine candidates. Patient recovery for those who are unvaccinated depends heavily on an early diagnosis and careful management of fever symptoms.
1Only female mosquitoes bite and drink blood. Female mosquitoes need the protein in blood to make their eggs.
2See http://www.who.int/immunization/research/development/dengue_vaccines/en [accessed May 2, 2016].
TABLE 3-1 Selected Case Studies on Gene Drive Research and Related Applications
|1||Use the mosquitoes Aedes aegypti and Aedes albopictus to manage dengue||
Credit: James Gathany
|2||Using Anopheles gambiae mosquitoes to combat malaria||
Credit: James Gathany
|3||Using the Culex quinquefasciatus mosquitoes to combat avian malaria||
Mosquito biting a honeycreeper
Credit: Susanne Bard
|4||Controlling populations of non-indigenous Mus musculus mice to protect native biodiversity on islands throughout the world||
|5||Controlling non-indigenous Centaurea maculosa knapweeds to protect biodiversity in rangelands and forests in the United States||
Credit: Matt Levin
|6||Controlling Amaranthus palmeri (Palmer amaranth, also known as pigweed) to increase agriculture productivity in the Southern United States||
Credit:Michael J. Plagens
|7||Developing a vertebrate model for gene drive research using Danio rerio zebrafish||
Credit: Monte Westerfield
Current Mitigation Efforts
Prevention of dengue relies entirely on vector control, mostly through ultra-low volume spraying of insecticides. Insecticide resistance is challenging the efficacy of such dengue vector control methods using currently available chemicals. Another vector control intervention is the
management of mosquito vector breeding sites, which are typically man-made containers. However, because dengue disease exhibits spatiotemporal heterogeneity epidemic activity (alternating as high and low incidences between years and seasons), and because of the potential serotype interaction and co-circulations, predicting possible epidemics is extremely complex as is effective prevention. These strategies are laborious and typically reactive rather than proactive (Achee et al., 2015). Additional control strategies are listed in Appendix C.
Biological controls also exist, such as the use of cyclopoid copepods (Marten et al., 1994), population reduction via community participation (Scholte et al., 2006; Majambere et al., 2007) and the use of larvivorous fish, but the maintenance of the distributed containers is a limiting factor to effective control. Another type of biological control is through the release of Wolbachia-infected mosquitoes. The bacterial symbionts in the genus Wolbachia are widely distributed in insects (Werren et al., 1995; Werren and O’Neil, 1997; Bourtzis and Braig, 1999; Stouthamer et al., 1999). Wolbachia infection reduces the lifespan of the insect hosts (Sinkins et al., 1997; Dobson et al., 2002; Ahantarig et al., 2011; Bull and Turelli, 2013). In addition, Wolbachia infection of Aedes aegypti confers resistance to infection with dengue and chikungunya viruses (McMeniman et al., 2009; Moreira et al., 2009; Bian et al., 2010). In light of these results, small-scale trials to reduce dengue transmission using Wolbachia started in 2011 in Australia and further expanded to Vietnam, Indonesia, and Brazil.3 Although on-going large field trials suggest a reduction of dengue incidence, there remain important considerations concerning the unanticipated evolution of the dengue virus (or other viruses infecting the same mosquito vector) that need to be addressed.
In summary, despite many available methods of mosquito control, existing methods are not yet fully effective at reducing dengue transmission.
Plausibility of a Gene Drive Solution
It may be possible to create two types of gene drives in Aedes species: one that prevents the transmission of the dengue virus and another that causes sterility. Research with Wolbachia demonstrates, in principle, the potential for those two approaches. In 2010, researchers showed that Wolbachia can be used to induce resistance in Aedes aegypti to the dengue virus. Wolbachia also can be used to shorten the life-span of Aedes aegypti (McMeniman et al., 2009). Similarly, the U.K.-based company Oxitec has developed a technology to suppress Aedes aegypti populations in which male Aedes aegypti mosquitoes are genetically engineered to be sterile.4 The first proofs-of-concept experiments demonstrating the creation of a gene drive in the fruit fly, a model organism for invertebrate research, and in other mosquito species (discussed below) also provide evidence that a gene drive could be developed in Aedes aegypti (Gantz and Bier, 2015; Gantz et al., 2015; Hammond et al., 2016). These applications would require initial release of a number of the gene-drive modified mosquitoes within an urban setting where dengue is endemic or where dengue outbreaks are known.
Create gene drives in Anopheles gambiae mosquitoes to reduce the spread of human malaria in sub-Saharan Africa.
Malaria is a serious and sometimes fatal parasitic infection that occurs in nearly 100 countries worldwide. Adults and children who contract malaria often experience high fever and anemia. If the infection is severe, coma and death can occur. Malaria disproportionately affects people, particularly children, in low and middle income countries in sub-Saharan Africa, South Asia, and South America.
Human malaria is caused by any of the five protozoan parasites of the Plasmodium genus. The mosquito Anopheles gambiae is the primary vector (carrier) of Plasmodium in sub-Saharan Africa.
Current Mitigation Efforts
Current methods for malaria control focus on two themes, drug therapy and vector control. The ability to treat infection requires detection of the parasite and access of infected persons to healthcare, which can be extremely challenging in many, if not most, malaria-endemic settings. Malaria vaccines are under development and have shown promise, but will take many more years before they can be fully recommended for wide application. Prevention of transmission targeting the Anopheline mosquito vector is based on interventions recommended by the World Health Organization. These include measures to eliminate breeding sites, spraying insecticides with residual properties onto the walls of houses, and using insecticide-treated bed nets in areas where malaria is endemic. Additional control strategies are listed in Appendix C. However, all of these measures require organized campaigns and sustained resource availability. In addition, efforts to control malaria are in jeopardy due to the spread of insecticide resistance in Anopheles gambiae populations (Edi et al., 2012; Namountougou et al., 2012; Cisse et al., 2015).
Plausibility of a Gene Drive Solution
A gene drive that alters the female mosquito’s ability to become infected with the malaria parasite, or one that prevents parasite development within the mosquito, could block malarial transmission without affecting mosquito populations. In November 2015, researchers demonstrated that CRISPR/Cas9 can be used to create a gene drive that could spread anti-Plasmodium genes in populations of a malaria-carrying Anopheline mosquito, Anopheles stephensi (Gantz et al., 2015). However, the system transmits the drive construct at Mendelian frequencies in some crosses, suggesting that this valuable proof-of-principle needs further modification and research before field release (Gantz et al., 2015). Alternatively, a gene drive that alters the fitness of the female mosquito could result in reducing vector populations over time. In December 2015, researchers demonstrated that CRISPR/Cas9 can be used to create a gene drive that causes sterility in female Anopheles gambiae mosquitoes (Hammond et al., 2016). Although one of the research team’s constructs is predicted to spread through a population, it has not yet been shown to spread to high frequency in a population containing heterogeneous genetic backgrounds. Nonetheless, the anti-Plasmodium and the female sterility gene drive approaches theoretically have the potential to eliminate malaria in sub-Saharan African villages where malaria is endemic.
Create gene drives in southern house mosquitoes, Culex quinquefasciatus, to reduce the spread of avian malaria to threatened and endangered honeycreeper birds in the Hawaiian Islands.
Avian malaria is a disease caused by protozoan parasites that infect birds. Birds become infected when they are “bitten” by female mosquitoes carrying the parasite. Birds without immune resistance to the parasite become anemic, grow progressively weaker, and ultimately die. Avian malaria is common in most continents, but absent from many isolated islands where mosquitoes (and hence Plasmodium) do not naturally occur (Atkinskon, 2005).5 Thus, native birds in Hawaii, the Galapagos, and other archipelagoes, which evolved without natural exposure to Plasmodium parasites, are highly susceptible to avian malaria. The southern house mosquito, Culex quinquefasciatus, is the primary mosquito vector of Plasmodium relictum in Hawaii. The displacement and extinction of native birds has greatly impacted ecological systems and biodiversity in Hawaii, and climate change threatens to expand mosquito ranges into higher elevations, thereby presenting greater harm to bird populations at these elevations.
Current Mitigation Efforts
Prevention of avian malaria transmission has historically been through interventions that target mosquito vector populations using insecticide spraying and larval source management. Similar to resistance of parasites to drugs, many mosquito species are resistant to currently available chemicals, making control difficult. In Hawaii, attempts to control the mosquitoes through such methods have not eliminated the threat. See Appendix C for a comprehensive list of mosquito control strategies.
Plausibility of a Gene Drive Solution
The use of gene drives could be used as a new strategy to target the mosquito vector to control avian malaria. As described in the first two case studies, there is strong potential to develop gene drives that alter the female mosquito’s ability to become infected with the malaria parasite, or that prevent mosquitoes from reproducing. The first proofs-of-concepts in which gene drives were created in the fruit fly and in other mosquito species provide evidence that a gene drive could also be developed in Culex quinquefasciatus (Gantz and Bier, 2015; Gantz et al., 2015; Hammond et al., 2016).
CASE STUDY 4: CONTROLLING POPULATIONS OF NON-INDIGENOUS MUS MUSCULUS MICE TO PROTECT BIODIVERSITY ON ISLANDS
Reduce or eliminate populations of the non-indigenous mouse, Mus musculus, to protect native biodiversity on islands around the world.
Invasive species are a leading cause of extinction of native wildlife and plants on islands. Nearly half of all species included on the International Union for the Conservation of Nature’s list of species that are threatened with extinction live on islands. In addition, roughly 70%, 90%, and 95% of all extinctions of mammals, reptiles, and birds occur on islands, respectively (Campbell et al., 2015; Godwin, 2015). The activities of the house mouse, Mus musculus, and other
introduced rodents reduce the ability of native species to reproduce, alter or destroy habitats so that they no longer support the needs of native species, and in other ways negatively affect island ecosystem dynamics. Approximately 80% of the world’s islands now have invasive rodents (Campbell et al., 2015; Godwin, 2015).
Current Mitigation Efforts
Efforts to eradicate rodents from islands include the use of traps, poisons, and biological controls, such as the introduction of predators or diseases. Application of rodenticides can be cost-prohibitive due to expenses associated with regulation compliance, dispersal method, size of the treated area, and cost of the toxicant itself (Meerburg et al., 2008; Williams, 2013). Mechanical traps are often considered more humane than rodenticides because they do not involve the use of chemicals that could adversely affect human, animal, and overall ecosystem health (Lorvelec and Pascal, 2005; Witmer et al., 2011). However, placing traps and collecting the caught animals is labor intensive, traps do not discriminate between target and non-target organisms (Lorvelec and Pascal, 2005), and traps are insufficient to fully eradicate a rodent population without the use of other methods. Other research aims to use genetic engineering approaches to control rodent populations including RNA interference and developing transgenes that cause female progeny to develop as males or prevent all progeny from developing (Gemmell et al., 2013; He et al., 2015). It remains to be seen if such genetic engineering approaches will be effective, scalable and affordable (Jacob et al., 2008; Campbell et al., 2015). Additional discussion of these methodologies and a more comprehensive list of other approaches used to control rodent populations are presented in Appendix D.
Plausibility of a Gene Drive Solution
Scientists are studying a sex-determining gene drive that causes house mice to produce more male offspring than females (Cocquet et al., 2012). If this occurs over multiple generations, it should lead to a reduction in population size over time. The molecular mechanism takes advantage of an endogenous region of high meiotic drive (meaning it is more likely to be inherited) in the mouse genome found on chromosome 17 (an autosome) called the t-complex. In this scenario, male mice are genetically engineered to possess the Sry gene, which promotes male characteristics (Goodfellow and Lovell-Badge, 1993), on chromosome 17 instead of its usual location on the Y chromosome. An XY Sry male is fertile, and upon mating to a wild-type XX female, both the XY and XX offspring (both male and females) possess Sry and physically develop into male mice, with XX male mice being sterile and the XY mice still able to reproduce and transmit Sry. Over time, the population of mice would tend to become all male, leading to a decrease in reproduction and eventual population decline and suppression due to the loss of female mice (Campbell et al., 2015). Male mice are promiscuous, and so have nearly an unlimited amount of reproductive potential, as long as fertile female mice are present. Female mice must go through a gestation period after mating, limiting their ability to contribute their genetic information to future generations. Hence, female mice are the limiting factor in the change of population densities over time. A description of the technique, and elements that helped in the development of a case study in this report can be found on a website dedicated to island conservation created by students from North Carolina State University.6
Other potential gene drive mechanisms based upon Medea or underdominance strategies could also be used to achieve the same purpose and would involve inducing targeted translocations into the mouse genome.
CASE STUDY 5: CONTROLLING NON-INDIGENOUS CENTAUREA MACULOSA KNAPWEEDS TO PROTECT BIODIVERSITY IN RANGELANDS AND FORESTS
Create gene drives in the non-indigenous knapweed species, Centaurea maculosa, to protect biodiversity of native plant species in rangelands and forests in the United States.
The spotted knapweed (Centaurea maculosa) is native to Eastern Europe but was introduced to the United States in the late 1800s. By the year 2000, spotted knapweed could be found in 45 of the 50 states and covered nearly 7 million acres of rangeland and pine forest (Zouhar, 2001). Spotted knapweed first invades disturbed habitats; once established, it spreads to native ecosystems, causing soil erosion in the process.
Current Mitigation Efforts
Several attempts have been made to slow the spread of spotted knapweed by using biological controls; these reduce seed production but have not had large effects on the density of Centaurea maculosa plants (Sheley et al., 1998). In addition to biological controls, management of knapweed populations has focused on physical removal, fire, and chemical treatment for infestations (Sheley et al., 1998; Zouhar, 2001).
Plausibility of a Gene Drive Solution
Spotted knapweed is obligately outcrossing (Harrod and Taylor, 1995), meaning that there is little or no self-fertilization and that gene drives would be able to spread throughout knapweed populations. Another factor that makes it potentially suitable for a gene drive is that the basis for its ability to outcompete native plants is thought to come from the production of a compound called catechin (Thelen et al., 2005), which it exudes from its the roots. Catechin inhibits the germination and growth of native plant species, thereby conferring a competitive advantage to spotted knapweed (Bais et al., 2003).
There are two possible gene drive approaches to help limit the spread of spotted knapweed, which could potentially be employed together. The first option is to engineer a suppression gene drive by targeting sex-specific genes, thereby biasing gender ratios and facilitating a population crash. The second is to modify the population by targeting the catechin biosynthetic pathway, which in theory would negatively affect the knapweed’s ability to compete against endemic plants, although this effect is still debated (Perry et al., 2005). In either case, the rate of spread of either of these gene drives is expected to be slow, because spotted knapweed is a perennial plant that lives for approximately 9 years (Zouhar, 2001). In addition, the success of a suppression drive is likely to depend critically on the fertility advantages of sex-modified plants compared to hermaphrodites and also on features such as pollen availability and spatial structure (Hodgins et al., 2008).
Retraction: In March 2016, the Journal of Ecology and authors Laura G. Perry, Ragan Callaway, and Jorge Vivanco retracted a research publication on the ability of knapweed to outcompete native plants through the production of catechin. Given the retraction, a gene drive that inhibits catechin production in knapweed, as discussed in Case Study 5 on page 56 of this report, is no longer considered plausible. The retraction does not affect discussions of Case Study 5 in other chapters, or the reportâ€™s conclusions and recommendations. Details about the retraction are accessible online at: http://onlinelibrary.wiley.com/doi/10.1111/1365-2745.12560/full.
Create gene drives in Palmer amaranth (Amaranthus palmeri also called pigweed), to reduce or eliminate the weed on agricultural fields in the Southern United States.
Palmer amaranth infests agricultural fields throughout the American South. It has evolved resistance to the herbicide glyphosate, the world’s most-used herbicide (Powles, 2008), and this resistance has become geographically widespread.
Current Mitigation Efforts
Whether a plant is considered a weed is context-dependent. In one region, a plant is desirable, whereas in another place, the same plant may be a weed. A plant is typically viewed as a “weed” when it has little recognized value in the locale where it is growing and when it grows rapidly and competes with a crop or pastureland for space, light, water, and nutrients. Weed management is a continual and major challenge. In addition to competition for resources and interfering with the management of desirable plants, poisonous weeds can negatively impact human health, crops and livestock (Bridges et al., 1994). Management strategies fall into four major categories: physical and mechanical methods, cultural methods, chemical methods, and biological methods. Examples of mechanical practices include manual removal of weeds, which is labor intensive, or tilling, which can increase soil erosion. Examples of cultural practices include crop rotations using plants that choke out weeds (often there are limited choices available) and using drip irrigation to limit water to planting rows, which only works well in dry regions that extensively irrigate. Examples of biological methods include animal grazing and the use of natural enemies (microbes, insects, and other animals such as nematodes, fish, and birds); these strategies are primarily used in low-intensity management of rangelands, forests, preserved natural areas, and waterways.
In much of production agriculture, the primary approach to control weeds is to use herbicides. Glyphosate, the most commonly used herbicide, is a systemic herbicide that, when applied, moves throughout the plant thus destroying more tissues as compared to contact herbicides. The generation of herbicide-resistant crops has revolutionized weed control. Glyphosate-resistant crops have been rapidly adopted in multiple crops because of economic advantages, strong weed control, and the observation that the glyphosate-resistant crop system confers a lower environmental impact than the approaches it replaced (Duke and Powles, 2009). Unfortunately, after decades of glycophosate use weeds are now adapting, and herbicide resistance is increasing among weed population, reducing the efficacy of glyphosate for weed control (Powles and Yu, 2010). The current strategy to deal with herbicide-resistant weeds is to adopt diverse tactics, combining multiple weed control approaches (Duke and Powles, 2009; Norsworthy et al., 2012). The particular combinations of strategies chosen depend on the crop, the region, and the major weeds impacting the particular agricultural system. Details on specific practices can be found on agricultural extension websites at land grant institutions throughout the United States and at equivalent international institutions’ websites.
Plausibility of a Gene Drive Solution
Palmer amaranth is a likely candidate for gene drive technology, for five reasons. First, it is an annual plant, so it has yearly sexual reproduction and a rapid generation time. Second, Palmer amaranth and some other members of the genus are dioecious (male and female flowers occur on separate plants) (Steckel, 2007), which ensures the outcrossing necessary to spread gene drives.
Third, it does not have an extensive seed bank; studies suggest that most seeds do not persist in the soil, so that there is unlikely to be a seed repository that is immune to the gene drive. Fourth, an Amaranthus species has been transformed genetically (Pal et al., 2013), suggesting that it will be technologically feasible to insert gene drives into Palmer amaranth. Finally, Palmer amaranth is wind-pollinated, implying that the eradication of species will, at the very least, not harm insect pollinators.
In theory, Palmer amaranth could be subjected to two types of gene drive. In the first, a modification drive would target the genes that confer resistance to glyphosate and reestablish the population’s susceptibility to glycophosate herbicides. The potential targets of this gene drive are known, because the glyphosate herbicide acts by interrupting the function of 5-enolpyruvylshikimate-3-phosphate synthase. In Palmer amaranth, this synthase gene has been duplicated extensively, leading to enzyme overexpression and glyphosate resistance (Gaines et al., 2010). Thus, a candidate gene drive would need to target multiple 5-enolpyruvylshikimate-3-phosphate synthase copies that are scattered throughout the genome. If the gene drive succeeded and susceptibility became fixed, glyphosate could then be used again as a tool to limit Palmer amaranth populations.
A second approach would be to build a suppression drive. Although the target and content of such a drive is not yet clear, the fact that there are separate male and female plants implies that there are sex-specific genes that are suitable targets for biasing the sex ratio. Under this approach, the goal would be skew sex ratios until the entire population (or species) collapses.
Create gene drives in the zebrafish, Danio rerio, to study gene drive mechanisms in a vertebrate animal.
As of April 2016, researchers have not developed a gene-drive modified vertebrate for use in fundamental research in the laboratory but proofs-of-concept for gene drives have been demonstrated in yeast, the fruit fly, and mosquitoes, with the expectation that this technique will be translated to a vertebrate animal at a future date (DiCarlo et al., 2015; Gantz and Bier, 2015; Gantz et al., 2015; Hammond et al., 2016). These current animal models, and the behavior of gene drives in them, will not necessarily recapitulate the behavior of gene drives in vertebrate species. Given the fundamental differences between vertebrates and invertebrates, a vertebrate species for gene drive research will be needed to address a variety of fundamental research topics before using gene drives in other vertebrate animals, particularly those intended for release into the environment; and also potentially to make comparisons with gene drive mechanisms in invertebrates.
Current Mitigation Efforts
7A mouse could also potentially be a candidate vertebrate model for gene drive research. Research on the naturally occurring t-complex in mice offers insight into how regions of high meiotic drive function and affect characteristics associated with vertebrate development and behavior (see Case Study 4). However, these studies may not be broadly applicable to other vertebrates. Also, the gestation period, and thus the generation time, is longer in mice than in zebrafish, which could make it more difficult for research to keep pace with rapid advances in invertebrates. However, existing approaches for gene editing through transient introduction of CRISPR/Cas9 (or other mechanisms) have been successful; thus, the committee considers development of a gene-drive modified mouse for laboratory research plausible, a close second to the case study on zebrafish presented in this report.
Containment of zebrafish is straightforward due to the requirement for appropriate aquatic facilities, while other potential vertebrate models for gene drives, such as the mouse, could more easily escape from, and survive outside, the laboratory. In addition, it may be possible to develop
a self-limiting gene drive in zebrafish by making the drive active only in the presence of tetracycline, which could be required to activate the promoter needed to express the gene drive construct (Hammond et al., 2016).
Plausibility of a Gene Drive Solution
A gene-drive modified zebrafish could be developed specifically for laboratory studies with no intention for environmental release. The zebrafish provides an outstanding model to address basic research questions about gene drives in a vertebrate species for many reasons (Shah and Moens, 2016). The zebrafish genome has been fully sequenced, and zebrafish have well-characterized traits associated with reproduction and other behaviors (Howe et al., 2013). Zebrafish are also low cost and easy to maintain, have a short generation time, and produce large numbers of offspring (Lawrence et al., 2012; Harris et al., 2014). They are also preferred from a regulatory standpoint (e.g., from the standpoint of Institutional Animal Care and Use Committee) with regards to using animal models for research. Moreover, gene editing has already been used successfully in this organism (Ma and Liu, 2015; D’Agostino et al., 2016; Lin et al., 2016; Prykhozhij et al., 2016).
A gene-drive modified zebrafish could be created by inserting a gene drive construct into the fish consisting of Cas9, a gRNA targeting a non-essential locus (e.g., a gene expressed in the eye) and a green fluorescent protein marker to identify the gene-drive modified organism. The latter characteristic would give rise to a visible phenotype upon insertion of the donor template on the construct.
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