A CRISPR–dCas Toolbox for Genetic Engineering and Synthetic Biology

Xu, Xiaoshu; Qi, Lei S.

doi:10.1016/j.jmb.2018.06.037

Cited by 247 publications

(169 citation statements)

References 119 publications

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“…These constructs are designed from a general standpoint and seek to translate each of the involved components using transcriptional logic gates, intercellular communication mechanisms, and external elements such as environmental signals. It should be highlighted that the mapping presented here is a proposal and could be complemented and extended with other kinds of mechanisms, such as CRISPR 19,20 systems, external conditions such as temperature, nutrient consumption, or specific spatial conditions. Also, it should be stressed that our proposal heavily relies on intercellular communication, since it offers a higher computation power and also distributes it among the colony cells.…”

Section: Resultsmentioning

confidence: 99%

A framework for implementing metaheuristic algorithms using intercellular communication

Gutiérrez

Ortiz

Carrión

2020

Preprint

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Metaheuristic procedures (MH) have been a trend driving Artificial Intelligence (AI) researchers for the past 50 years. A variety of tools and applications (not only in Computer Science) stem from these techniques. Also, MH frequently rely on evolution, a trademark process involved in cell colony growth. Generally, MH are used to approximate the solution to difficult problems but require a large amount of computational resources. Cell colonies harboring synthetic distributed circuits using intercell communication offer a direction for tackling this problem, as they process information in a massively parallel fashion. In this work, we propose a framework that maps MH elements to synthetic circuits in growing cell colonies. The framework relies on cell-cell communication mechanisms such as quorum sensing (QS) and bacterial conjugation. As a proofof-concept, we also implemented the workflow associated to the framework, and tested the execution of two specific MH (Genetic Algorithms and Simulated Annealing) encoded as synthetic 2 circuits on the gro simulator. Furthermore, we show an example of how our framework can be extended by implementing another kind of computational model: The Cellular Automaton. This work seeks to lay the foundations of mappings for implementing AI algorithms in a general manner using Synthetic Biology constructs in cell colonies. MAIN TEXTEvolution is a key element involved in all microbiology processes. It is the process that drives organism adaptation to better survive and thrive in their surrounding environment. This process occurs at a genetic level, involving mainly genetic recombination and mutation. The genetic diversity produced by evolution is studied and used as inspiration in computational methods such as Evolutionary Algorithms (EAs) 1,2 . These algorithms are generally used for approximating solutions to optimization problems. Since evolution is a standard occurring process, it is natural to relate EAs to microbiology experiments, and more specifically, to Synthetic Biology constructs. This relationship has already been addressed by Directed Evolution 3,4 . However, the control level of Directed Evolution is not as specific as the one reached in EAs. Furthermore, several other computational methods can be translated to Synthetic Biology constructs that emulate their operation. Metaheuristic procedures (MH) 5-7 are a larger class of procedures that contain EAs.Inspiration upon which these techniques are designed range from metallurgy processes 8-10 through bird flock movement patterns [11][12][13] , and ant colony food foraging 14,15 . A general mapping, relating Synthetic Biology constructs to MH elements can be proposed such that any procedure of that class can be modeled as a synthetic circuit. This is due to MH sharing common elements and similarities that can be generalized.

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Section: Resultsmentioning

confidence: 99%

A framework for implementing metaheuristic algorithms using intercellular communication

Gutiérrez

Ortiz

Carrión

2020

Preprint

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“…Programmable dCas9‐based systems offer increasingly diverse tools for investigations of gene function in higher plants. By mutating the catalytic domains of Cas9, the deactivated Cas9 (dCas9) can be fused to a variety of functional domains and then targeted to a specific genomic region by the gRNA (Xu et al ). For example, dCas9‐VP64 and dCas9‐SRDX were respectively shown to activate or repress endogenous genes in Arabidopsis and tobacco (Lowder et al ; Lowder et al ) (Figure A, B).…”

Section: Discussionmentioning

confidence: 99%

“…A successful application was reported in creating herbicide‐resistant watermelon as described earlier (Tian et al ). In addition, dCas9 can be fused to chromatin modifying enzymes including DNA methytransferase, histone deacetylase, and others to epigenetically modulate transcription at specific genomic locus (Xu et al ) (Figure D).…”

Section: Discussionmentioning

confidence: 99%

Application and future perspective of CRISPR/Cas9 genome editing in fruit crops

Zhou¹,

Wang

et al. 2019

JIPB

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Fruit crops, including apple, orange, grape, banana, strawberry, watermelon, kiwifruit and tomato, not only provide essential nutrients for human life but also contribute to the major agricultural output and economic growth of many countries and regions in the world. Recent advancements in genome editing provides an unprecedented opportunity for the genetic improvement of these agronomically important fruit crops. Here, we summarize recent reports of applying CRISPR/Cas9 to fruit crops, including efforts to reduce disease susceptibility, change plant architecture or flower morphology, improve fruit quality traits, and increase fruit yield. We discuss challenges facing fruit crops as well as new improvements and platforms that could be used to facilitate genome editing in fruit crops, including dCas9‐base‐editing to introduce desirable alleles and heat treatment to increase editing efficiency. In addition, we highlight what we see as potentially revolutionary development ranging from transgene‐free genome editing to de novo domestication of wild relatives. Without doubt, we now see only the beginning of what will eventually be possible with the use of the CRISPR/Cas9 toolkit. Efforts to communicate with the public and an emphasis on the manipulation of consumer‐friendly traits will be critical to facilitate public acceptance of genetically engineered fruits with this new technology.

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“…The Cas13a-CRISPRi has certain advantages over the dCas9-CRISPRi for repressing gene expression 16,34 . First, the dCas9-CRISPRi machinery acts at DNA level while Cas13a targets mRNA directly.…”

Section: Discussionmentioning

confidence: 99%

Programmable CRISPR interference for gene silencing using Cas13a in mosquitoes

Kulkarni

Moon

et al. 2019

Preprint

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In the CRISPR-Cas systems, Cas13a is an RNA-guided RNA nuclease specifically targeting single strand RNA. In this study we developed a Cas13a mediated CRISPR interference tool to target mRNA for gene silencing in mosquitoes and tested the machinery in two mosquito species. A Cas13a expressing plasmid was delivered to mosquitoes by intrathoracic injection, and Cas13a transcripts were detectable at least 10 days post-delivery. In Anopheles gambiae, vitellogenin (Vg) gene was silenced by Vg-crRNA injection two hours post blood meal, which was accompanied by a significant reduction in egg production. In Aedes aegypti, the α -and δ subunits of COPI genes were knocked down by a post-blood meal crRNA injection, which resulted in mortality and fragile midguts, reproducing a phenotype reported in a RNAi mediated COPI gene silencing study. The silencing of mulitple genes simlutaneously is achievable when mosquitoes are given a cocktail of Cas13a construct with crRNAs targeting respective genes.This study adds a programable CRISPR tool to manipulate RNA in insects.

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A CRISPR–dCas Toolbox for Genetic Engineering and Synthetic Biology

Cited by 247 publications

References 119 publications

A framework for implementing metaheuristic algorithms using intercellular communication

A framework for implementing metaheuristic algorithms using intercellular communication

Application and future perspective of CRISPR/Cas9 genome editing in fruit crops

Programmable CRISPR interference for gene silencing using Cas13a in mosquitoes

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