scholarly journals Genetic conversion of a split-drive into a full-drive element

2021 ◽  
Author(s):  
Gerard Terradas ◽  
Jared B. Bennett ◽  
Zhiqian Li ◽  
John M. Marshall ◽  
Ethan Bier
Keyword(s):  
Gene Drive ◽  
Fitness Costs ◽  
Experimental Proof ◽  
Guide Rna ◽  
Split Gene ◽  
Core Components ◽  
Beneficial Traits ◽  
Drive Systems ◽  
Genetic Conversion ◽  
Gene Drives

AbstractGene-drive systems offer an important new avenue for spreading beneficial traits into wild populations. Their core components, Cas9 and guide RNA (gRNA), can either be linked within a single cassette (full gene drive, fGD) or provided in two separate elements (split gene drive, sGD) wherein the gRNA-bearing element drives in the presence of an independent static source of Cas9. We previously designed a system engineered to turn split into full gene drives. Here, we provide experimental proof-of-principle for such a convertible system inserted at the spo11 locus, which is recoded to restore gene function. In multigenerational cage studies, the reconstituted spo11 fGD cassette initially drives with slower kinetics than the unlinked sGD element (using the same Mendelian vasa-Cas9 source), but eventually reaches a similar level of final introgression. Different kinetic behaviors may result from transient fitness costs associated with individuals co-inheriting Cas9 and gRNA transgenes during the drive process.

scholarly journals Invasion and migration of spatially self-limiting gene drives: a comparative analysis

2017 ◽  
Author(s):  
Sumit Dhole ◽  
Michael R. Vella ◽  
Alun L. Loyd ◽  
Fred Gould
Keyword(s):  
Target Population ◽  
Gene Drive ◽  
Fitness Costs ◽  
Invasion And Migration ◽  
Migration Rates ◽  
Chain System ◽  
Drive Systems ◽  
And Migration ◽  
The One ◽  
Gene Drives

AbstractRecent advances in research on gene drives have produced genetic constructs that could theoretically spread a desired gene (payload) into all populations of a species, with a single release in one place. This attribute has advantages, but also comes with risks and ethical concerns. There has been a call for research on gene drive systems that are spatially and/or temporally self-limiting. Here we use a population genetics model to compare the expected characteristics of three spatially self-limiting gene drive systems: one-locus underdominance, two-locus underdominance, and daisy-chain drives. We find large differences between these gene drives in the minimum release size required for successfully driving a payload into a population. The daisy-chain system is the most efficient, requiring the smallest release, followed by the two-locus underdominance system, and then the one-locus underdominance system. However, when the target population exchanges migrants with a non-target population, the gene drives requiring smaller releases suffer from higher risks of unintended spread. For payloads that incur relatively low fitness costs (up to 30%), a simple daisy-chain drive is practically incapable of remaining localized, even with migration rates as low as 0.5% per generation. The two-locus underdominance system can achieve localized spread under a broader range of migration rates and of payload fitness costs, while the one-locus underdominance system largely remains localized. We also find differences in the extent of population alteration and in the permanence of the alteration achieved by the three gene drives. The two-locus underdominance system does not always spread the payload to fixation, even after successful drive, while the daisy-chain system can, for a small set of parameter values, achieve a temporally-limited spread of the payload. These differences could affect the suitability of each gene drive for specific applications.Note:This manuscript has been accepted for publication in the journal Evolutionary Applications and is pending publication. We suggest that any reference to or quotation of this article should be made with this recognition.

scholarly journals Daisy-chain gene drives for the alteration of local populations

2019 ◽  
Vol 116 (17) ◽  
pp. 8275-8282 ◽  
Author(s):  
Charleston Noble ◽  
John Min ◽  
Jason Olejarz ◽  
Joanna Buchthal ◽  
Alejandro Chavez ◽  
...  
Keyword(s):  
Drive System ◽  
Chain Gene ◽  
Gene Drive ◽  
Rna Sequences ◽  
Guide Rna ◽  
Highly Active ◽  
Wide Range ◽  
Fitness Parameters ◽  
Drive Systems ◽  
Gene Drives

If they are able to spread in wild populations, CRISPR-based gene-drive elements would provide new ways to address ecological problems by altering the traits of wild organisms, but the potential for uncontrolled spread tremendously complicates ethical development and use. Here, we detail a self-exhausting form of CRISPR-based drive system comprising genetic elements arranged in a daisy chain such that each drives the next. “Daisy-drive” systems can locally duplicate any effect achievable by using an equivalent self-propagating drive system, but their capacity to spread is limited by the successive loss of nondriving elements from one end of the chain. Releasing daisy-drive organisms constituting a small fraction of the local wild population can drive a useful genetic element nearly to local fixation for a wide range of fitness parameters without self-propagating spread. We additionally report numerous highly active guide RNA sequences sharing minimal homology that may enable evolutionarily stable daisy drive as well as self-propagating CRISPR-based gene drive. Especially when combined with threshold dependence, daisy drives could simplify decision-making and promote ethical use by enabling local communities to decide whether, when, and how to alter local ecosystems.

scholarly journals Experiments confirm a dispersive phenotype associated with a natural gene drive system

2021 ◽  
Vol 8 (5) ◽  
Author(s):  
Jan-Niklas Runge ◽  
Anna K. Lindholm
Keyword(s):  
Sperm Competition ◽  
Local Density ◽  
House Mice ◽  
Gene Drive ◽  
Wild Type ◽  
Fitness Costs ◽  
Oestrus Cycle ◽  
Drive Systems ◽  
Average Body ◽  
Average Body Weight

Meiotic drivers are genetic entities that increase their own probability of being transmitted to offspring, usually to the detriment of the rest of the organism, thus ‘selfishly’ increasing their fitness. In many meiotic drive systems, driver-carrying males are less successful in sperm competition, which occurs when females mate with multiple males in one oestrus cycle (polyandry). How do drivers respond to this selection? An observational study found that house mice carrying the t haplotype, a meiotic driver, are more likely to disperse from dense populations. This could help the t avoid detrimental sperm competition, because density is associated with the frequency of polyandry. However, no controlled experiments have been conducted to test these findings. Here, we confirm that carriers of the t haplotype are more dispersive, but we do not find this to depend on the local density. t -carriers with above-average body weight were particularly more likely to disperse than wild-type mice. t -carrying mice were also more explorative but not more active than wild-type mice. These results add experimental support to the previous observational finding that the t haplotype affects the dispersal phenotype in house mice, which supports the hypothesis that dispersal reduces the fitness costs of the t .

scholarly journals Evolutionary dynamics of CRISPR gene drives

2016 ◽  
Author(s):  
Charleston Noble ◽  
Jason Olejarz ◽  
Kevin M. Esvelt ◽  
George M. Church ◽  
Martin A. Nowak
Keyword(s):  
Evolutionary Dynamics ◽  
Evolutionary Stability ◽  
Clear Understanding ◽  
Gene Drive ◽  
Host Organism ◽  
Wild Populations ◽  
Selfish Genetic Elements ◽  
Real World Applications ◽  
Drive Systems ◽  
Gene Drives

AbstractThe alteration of wild populations has been discussed as a solution to a number of humanity’s most pressing ecological and public health concerns. Enabled by the recent revolution in genome editing, CRISPR gene drives, selfish genetic elements which can spread through populations even if they confer no advantage to their host organism, are rapidly emerging as the most promising approach. But before real-world applications are considered, it is imperative to develop a clear understanding of the outcomes of drive release in nature. Toward this aim, we mathematically study the evolutionary dynamics of CRISPR gene drives. We demonstrate that the emergence of drive-resistant alleles presents a major challenge to previously reported constructs, and we show that an alternative design which selects against resistant alleles greatly improves evolutionary stability. We discuss all results in the context of CRISPR technology and provide insights which inform the engineering of practical gene drive systems.

scholarly journals Dodging silver bullets: good CRISPR gene-drive design is critical for eradicating exotic vertebrates

2017 ◽  
Vol 284 (1860) ◽  
pp. 20170799 ◽  
Author(s):  
Thomas A. A. Prowse ◽  
Phillip Cassey ◽  
Joshua V. Ross ◽  
Chandran Pfitzner ◽  
Talia A. Wittmann ◽  
...  
Keyword(s):  
Female Sterility ◽  
Gene Drive ◽  
End Joining ◽  
Guide Rna ◽  
Deleterious Alleles ◽  
Precise Knowledge ◽  
Animal Populations ◽  
Individual Based Models ◽  
Non Homologous End Joining ◽  
Gene Drives

Self-replicating gene drives that can spread deleterious alleles through animal populations have been promoted as a much needed but controversial ‘silver bullet’ for controlling invasive alien species. Homing-based drives comprise an endonuclease and a guide RNA (gRNA) that are replicated during meiosis via homologous recombination. However, their efficacy for controlling wild populations is threatened by inherent polymorphic resistance and the creation of resistance alleles via non-homologous end-joining (NHEJ)-mediated DNA repair. We used stochastic individual-based models to identify realistic gene-drive strategies capable of eradicating vertebrate pest populations (mice, rats and rabbits) on islands. One popular strategy, a sex-reversing drive that converts heterozygous females into sterile males, failed to spread and required the ongoing deployment of gene-drive carriers to achieve eradication. Under alternative strategies, multiplexed gRNAs could overcome inherent polymorphic resistance and were required for eradication success even when the probability of NHEJ was low. Strategies causing homozygotic embryonic non-viability or homozygotic female sterility produced high probabilities of eradication and were robust to NHEJ-mediated deletion of the DNA sequence between multiplexed endonuclease recognition sites. The latter two strategies also purged the gene drive when eradication failed, therefore posing lower long-term risk should animals escape beyond target islands. Multiplexing gRNAs will be necessary if this technology is to be useful for insular extirpation attempts; however, precise knowledge of homing rates will be required to design low-risk gene drives with high probabilities of eradication success.

scholarly journals Synthetically EngineeredMedeaGene Drive System in the Worldwide Crop Pest,D. suzukii

2017 ◽  
Author(s):  
Anna Buchman ◽  
John M. Marshall ◽  
Dennis Ostrovski ◽  
Ting Yang ◽  
Omar S. Akbari
Keyword(s):  
Wild Population ◽  
Mendelian Inheritance ◽  
Gene Drive ◽  
Fitness Costs ◽  
High Frequencies ◽  
Toxin Resistance ◽  
Crop Pest ◽  
Naturally Occurring ◽  
Drive Systems ◽  
Significant Disease

AbstractSynthetic gene drive systems possess enormous potential to replace, alter, or suppress wild populations of significant disease vectors and crop pests; however, their utility in diverse populations remains to be demonstrated. Here, we report the creation of the first-ever syntheticMedeagene drive element in a major worldwide crop pest,D. suzukii. We demonstrate that this drive element, based on an engineered maternal “toxin” coupled with a linked embryonic “antidote,” is capable of biasing Mendelian inheritance rates with up to 100% efficiency. However, we find that drive resistance, resulting from naturally occurring genetic variation and associated fitness costs, can hinder the spread of such an element. Despite this, our results suggest that this element could maintain itself at high frequencies in a wild population, and spread to fixation, if either its fitness costs or toxin resistance were reduced, providing a clear path forward for developing future such systems.

scholarly journals Overcoming evolved resistance to population-suppressing homing-based gene drives

2016 ◽  
Author(s):  
John M. Marshall ◽  
Anna Buchman ◽  
Héctor M. Sánchez C. ◽  
Omar S. Akbari
Keyword(s):  
Mosquito Species ◽  
Gene Drive ◽  
Resistant Allele ◽  
Guide Rnas ◽  
Continental Scale ◽  
Fitness Advantage ◽  
Health Related ◽  
Drive Systems ◽  
Gene Drives

AbstractThe use of homing-based gene drive systems to modify or suppress wild populations of a given species has been proposed as a solution to a number of significant ecological and public health related problems, including the control of mosquito-borne diseases. The recent development of a CRISPR-Cas9-based homing system for the suppression ofAnopheles gambiae, the main African malaria vector, is encouraging for this approach; however, with current designs, the slow emergence of homing-resistant alleles is expected to result in suppressed populations rapidly rebounding, as homing-resistant alleles have a significant fitness advantage over functional, population-suppressing homing alleles. To explore this concern, we develop a mathematical model to estimate tolerable rates of homing-resistant allele generation to suppress a wild population of a given size. Our results suggest that, to achieve meaningful population suppression, tolerable rates of resistance allele generation are orders of magnitude smaller than those observed for current designs for CRISPR-Cas9-based homing systems. To remedy this, we propose a homing system architecture in which guide RNAs (gRNAs) are multiplexed, increasing the effective homing rate and decreasing the effective resistant allele generation rate. Modeling results suggest that the size of the population that can be suppressed increases exponentially with the number of multiplexed gRNAs and that, with six multiplexed gRNAs, a mosquito species could potentially be suppressed on a continental scale. We also demonstrate successful multiplexingin vivoinDrosophila melanogasterusing a ribozyme-gRNA-ribozyme (RGR) approach – a strategy that could readily be adapted to engineer stable, homing-based suppression drives in relevant organisms.

scholarly journals Analysis of off-target effects in CRISPR-based gene drives in the human malaria mosquito

2021 ◽  
Vol 118 (22) ◽  
pp. e2004838117
Author(s):  
William T. Garrood ◽  
Nace Kranjc ◽  
Karl Petri ◽  
Daniel Y. Kim ◽  
Jimmy A. Guo ◽  
...  
Keyword(s):  
Mendelian Inheritance ◽  
Gene Drive ◽  
Guide Rna ◽  
Human Malaria ◽  
Homology Directed Repair ◽  
Cas9 Nuclease ◽  
Set Up ◽  
Gene Drives ◽  
Target Effects ◽  
Important Milestone

CRISPR-Cas9 nuclease-based gene drives have been developed toward the aim of control of the human malaria vector Anopheles gambiae. Gene drives are based on an active source of Cas9 nuclease in the germline that promotes super-Mendelian inheritance of the transgene by homology-directed repair (“homing”). Understanding whether CRISPR-induced off-target mutations are generated in Anopheles mosquitoes is an important aspect of risk assessment before any potential field release of this technology. We compared the frequencies and the propensity of off-target events to occur in four different gene-drive strains, including a deliberately promiscuous set-up, using a nongermline restricted promoter for SpCas9 and a guide RNA with many closely related sites (two or more mismatches) across the mosquito genome. Under this scenario we observed off-target mutations at frequencies no greater than 1.42%. We witnessed no evidence that CRISPR-induced off-target mutations were able to accumulate (or drive) in a mosquito population, despite multiple generations’ exposure to the CRISPR-Cas9 nuclease construct. Furthermore, judicious design of the guide RNA used for homing of the CRISPR construct, combined with tight temporal constriction of Cas9 expression to the germline, rendered off-target mutations undetectable. The findings of this study represent an important milestone for the understanding and managing of CRISPR-Cas9 specificity in mosquitoes, and demonstrates that CRISPR off-target editing in the context of a mosquito gene drive can be reduced to minimal levels.

scholarly journals Recent advances in threshold-dependent gene drives for mosquitoes

2018 ◽  
Vol 46 (5) ◽  
pp. 1203-1212 ◽  
Author(s):  
Philip T. Leftwich ◽  
Matthew P. Edgington ◽  
Tim Harvey-Samuel ◽  
Leonela Z. Carabajal Paladino ◽  
Victoria C. Norman ◽  
...  
Keyword(s):  
High Frequency ◽  
Gene Drive ◽  
Fitness Costs ◽  
Practical Applications ◽  
Disease Vector ◽  
Recent Advances ◽  
Mathematical Analyses ◽  
Global Eradication ◽  
Gene Drives

Mosquito-borne diseases, such as malaria, dengue and chikungunya, cause morbidity and mortality around the world. Recent advances in gene drives have produced control methods that could theoretically modify all populations of a disease vector, from a single release, making whole species less able to transmit pathogens. This ability has caused both excitement, at the prospect of global eradication of mosquito-borne diseases, and concern around safeguards. Drive mechanisms that require individuals to be released at high frequency before genes will spread can therefore be desirable as they are potentially localised and reversible. These include underdominance-based strategies and use of the reproductive parasite Wolbachia. Here, we review recent advances in practical applications and mathematical analyses of these threshold-dependent gene drives with a focus on implementation in Aedes aegypti, highlighting their mechanisms and the role of fitness costs on introduction frequencies. Drawing on the parallels between these systems offers useful insights into practical, controlled application of localised drives, and allows us to assess the requirements needed for gene drive reversal.

scholarly journals Rodent gene drives for conservation: opportunities and data needs

2019 ◽  
Vol 286 (1914) ◽  
pp. 20191606 ◽  
Author(s):  
John Godwin ◽  
Megan Serr ◽  
S. Kathleen Barnhill-Dilling ◽  
Dimitri V. Blondel ◽  
Peter R. Brown ◽  
...  
Keyword(s):  
Food Security ◽  
Human Health ◽  
Genetic Model ◽  
Current Control ◽  
Invasive Pest ◽  
Gene Drive ◽  
Knowledge Gaps ◽  
Drive Systems ◽  
Invasive Rodents ◽  
Gene Drives

Invasive rodents impact biodiversity, human health and food security worldwide. The biodiversity impacts are particularly significant on islands, which are the primary sites of vertebrate extinctions and where we are reaching the limits of current control technologies. Gene drives may represent an effective approach to this challenge, but knowledge gaps remain in a number of areas. This paper is focused on what is currently known about natural and developing synthetic gene drive systems in mice, some key areas where key knowledge gaps exist, findings in a variety of disciplines relevant to those gaps and a brief consideration of how engagement at the regulatory, stakeholder and community levels can accompany and contribute to this effort. Our primary species focus is the house mouse, Mus musculus , as a genetic model system that is also an important invasive pest. Our primary application focus is the development of gene drive systems intended to reduce reproduction and potentially eliminate invasive rodents from islands. Gene drive technologies in rodents have the potential to produce significant benefits for biodiversity conservation, human health and food security. A broad-based, multidisciplinary approach is necessary to assess this potential in a transparent, effective and responsible manner.

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