Global ETD Search
Search theses and dissertations gathered from participating repositories worldwide. Every result links back to the library that holds it. No account is needed.
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Showing 1 to 19 of 19 for “"Cas12a"”.
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Methods for CRISPR Cas12a Multiplexing in Mammalian Systems
… mutants of CRISPR proteins such as Cas9 and Cas12a have allowed for a wide range of gene editing activities, including gene activation (CRISPRa). However, only a small number of genes can currently be targeted at one time, due to a limit on the number of gRNAs that can be successfully placed …
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The IN4MER CRISPR/Cas12a Multiplex Knockout Platform and Its Applications
… independent of background, and showed that the Cas12a platform exhibited enhanced sensitivity and consistency. Building on this knowledge, we developed a platform capable of expressing four independent enCas12a guide RNAs from a single promoter. Using this platform, we designed a whole-genome …
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Development of Universal Biosensing Platforms Based on CRISPR/Cas12a systems
… This thesis focusses on the development of CRISP/Cas12a based biosensing platforms for nucleic acid and protein detection. Two distinct CRISPR/Cas based diagnostic methods were developed. The first developed method is a plasmonic CRISPR Cas12a assay for colorimetric detection of viral nucleic …
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Genetic circuits for functional screens of Cas12a guide RNA libraries
… These variants hugely expand the toolbox of Cas12a components available to synthetic biologists, eliminating many of the current barriers to large-scale multiplexing.
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Utilizing the IN4MER CRISPR/CAS12A Multiplex Knockout Platform To Investigate Synthetic Lethality in the Human Genome
… candidate synthetic lethal paralogs; 2) our Cas12a multiplex platform “IN4MER” that provides superior sensitivity and replicability to support genome- wide and genetic interaction screening; 3) observations and analyses to account for varied guide efficiency in multiplex CRISPR screen …
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Mechanism of Permissive Cleavage Activity of TevCas12a
We created a novel dual endonuclease called TevCas12a. TevCas12a is a fusion of the I-TevI nuclease and linker domains to Cas12a. The goal was to create a dual endonuclease that made precise double-strand breaks leaving non-compatible ends for exogenous DNA insertion and specific deletions. …
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Development of a Multi-Component Biosensor for Recombinase Polymerase Amplification-CRISPR/Cas12a-Based Detection of Fusarium oxysporum f. sp. cubense tropical race 4
… Interspaced Short Palindromic Repeats (CRISPR)/Cas12a-mediated detection, and electrochemical square wave voltammetry (SWV) with targeted integration into a microfluidic system for amplification and detection within a compact heating unit. The research was organized into several key phases to …
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CRISPR-integrated Biosensors for Human Health: Novel strategies for pathogen detection and cancer screening
… in Escherichia coli (E. coli). Second, a CRISPR/Cas12a-based electrochemical sensor was designed for the detection of Salmonella Typhimurium in food samples. Third, a CRISPR/Cas14a system combined with blocker displacement amplification enabled sensitive detection of the BRAF V600E mutation in …
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Towards Accessible Diagnostics: Developing CRISPR-Based Colorimetric Tools for Low- and Middle-Income Countries
… and temperature versatility, CRISPR-Cas12a reactions were selected as the test platform. Design, production and screening of 48 gRNAs specific for S. Typhi were conducted considering various reaction parameters. The results were compared to existing gRNA prediction models. The …
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Developing Nucleic Acid-Based Sensors and Actuators
… materials using the CRISPR-associated nuclease Cas12a as a user-programmable sensor and material actuator. I exploit the programmability of Cas12a to actuate hydrogels containing DNA as an anchor for pendant groups or as a structural element. This versatile approach improves on the sensitivity …
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Engineering programmable nucleases for in vivo and in vitro applications
… editing. In Chapter 4, I introduced zCRISPR-Cas12a, an engineered CRISPR-Cas12a system that incorporates 2-aminoadenine (base Z) into the crRNA. This modification enhances the binding affinity between the crRNA and its complementary DNA target, resulting in a highly efficient and precise …
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Signal Processing Techniques Applied to Biomedical Diagnostics
… that pair this technology with isothermal CRISPR-Cas12a-based DNA/RNA amplification to detect clinically relevant targets with high specificity. Finally, we evaluate the clinical performance of our systems in detecting target genes for (1) SARS-CoV-2, the virus responsible for the COVID-19 …
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Mechanistic insight on a chimeric Cas9 protein's specificity for DNA target with 5 '-NAA-3' PAM
… our chimeric protein's comparative efficiency to Cas12a and the biophysical mechanism by which our grafted S. macacae segment works synergistically with the S. pyogenes Cas9 scaffold to cleave target DNA with an NAA PAM. We show that SpySmacCas9 does not bind or cleave at rates comparable to …
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Development and applications of programmable DNA-guided Argonaute-based artificial restriction enzymes
… method for cloning microbial BGCs named Cas12a assisted precise targeted cloning using in vivo Cre-lox recombination (CAPTURE). This method which consists of Cas12a digestion, a newly developed DNA assembly approach termed T4 polymerase exo + fill-in DNA assembly, and Cre-lox in vivo DNA …
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Impact of Manure Land Management Practices on Manure Borne Antibiotic Resistant Elements (AREs) in Agroecosystems
… ARGs (Spearman's ρ = 0.43-0.63). A CRIISPR-Cas12a assay for quantification of ARGs in environmental samples was just as precise as conventional methods. There is also potential for in-situ detection. These combined results can hopefully help farmers improve manure management practices that …
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NANOGP1 as a Model to Study the Consequences of Gene Duplications on Human Pluripotency and Development
… distinguish between the two genes. Using CRISPR/Cas12a-mediated gene editing in naïve PSCs, I introduced an epitope tag at the start of the predicted protein sequence, and this enabled me to demonstrate for the first time that endogenous NANOGP1 encodes an expressed protein. The ability to be …
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CRISPR-based approaches for the detection of antimicrobial-resistant genes
Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2025-05-01
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Multiplexed Crispr Libraries For Cancer Functional Genomics
<p>High-throughput forward genetic screenings are invaluable tools to systematically explore genetic interactions and to link gene disruption with disease contexts. The adaptation of CRISPR/Cas9 has improved the sensitivity and specificity of functional screenings. Despite this advance, there …