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 remains a long-standing need to improve functional screenings with smaller and more versatile pooled libraries. Capitalizing on the inherent multiplexing capability of a class 2 CRISPR enzyme AsCpf1, we developed a multiplexed, high throughput screening strategy that has avoided the usual trade-off between library size and library penetration, allowing library minimization without sacrificing gene targeting efficiency. We optimized the AsCpf1 protein for functional genomics use and demonstrated that an AsCpf1-based multiplexed library outperforms its monocistronic CRISPR/Cas9 library counterpart with a greatly reduced library size. With this strategy, we constructed the smallest whole-genome CRISPR knock-out library, Mini-human, for the human genome (n=17,032 constructs targeting 16,977 protein-coding genes), which performs favorably compared to conventional Cas9 libraries.
","abstract_html":"<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 remains a long-standing need to improve functional screenings with smaller and more versatile pooled libraries. Capitalizing on the inherent multiplexing capability of a class 2 CRISPR enzyme AsCpf1, we developed a multiplexed, high throughput screening strategy that has avoided the usual trade-off between library size and library penetration, allowing library minimization without sacrificing gene targeting efficiency. We optimized the AsCpf1 protein for functional genomics use and demonstrated that an AsCpf1-based multiplexed library outperforms its monocistronic CRISPR/Cas9 library counterpart with a greatly reduced library size. With this strategy, we constructed the smallest whole-genome CRISPR knock-out library, Mini-human, for the human genome (n=17,032 constructs targeting 16,977 protein-coding genes), which performs favorably compared to conventional Cas9 libraries.</p>","abstract_has_math":false,"creators":["Liu, Jintan","https://orcid.org/0000-0001-6609-9777
"],"institution":null,"degree_name":"Doctor of Philosophy (PhD)","degree_level":"Dissertation (PhD)","degree_discipline":null,"degree_department":null,"school":null,"contributors":["Giulio Draetta","Andy Futreal","Richard Behringer"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2020,"date_issued":"2020-05-01T07:00:00Z","date_published":"2020-05-01T07:00:00Z","updated_at":"2026-07-24T05:48:47Z","subjects":["CRISPR","Functional genomics","Pooled library screening","CRISPR/Cas12a","Biotechnology","Genomics","Medicine and Health Sciences"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://digitalcommons.library.tmc.edu/utgsbs_dissertations/995","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Giulio Draetta","Andy Futreal","Richard Behringer"]},{"key":"dc:creator","label":"Author","values":["Liu, Jintan","https://orcid.org/0000-0001-6609-9777
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"]},{"key":"dc:title","label":"Title","values":["Multiplexed Crispr Libraries For Cancer Functional Genomics"]}]}],"canonical_facts":{"dc:contributor":["Giulio Draetta","Andy Futreal","Richard Behringer"],"dc:creator":["Liu, Jintan","https://orcid.org/0000-0001-6609-9777
"],"dc:date.available":["2020-04-22T07:00:00Z"],"dc:description.abstract":["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 remains a long-standing need to improve functional screenings with smaller and more versatile pooled libraries. Capitalizing on the inherent multiplexing capability of a class 2 CRISPR enzyme AsCpf1, we developed a multiplexed, high throughput screening strategy that has avoided the usual trade-off between library size and library penetration, allowing library minimization without sacrificing gene targeting efficiency. We optimized the AsCpf1 protein for functional genomics use and demonstrated that an AsCpf1-based multiplexed library outperforms its monocistronic CRISPR/Cas9 library counterpart with a greatly reduced library size. With this strategy, we constructed the smallest whole-genome CRISPR knock-out library, Mini-human, for the human genome (n=17,032 constructs targeting 16,977 protein-coding genes), which performs favorably compared to conventional Cas9 libraries.
"],"dc:identifier":["https://digitalcommons.library.tmc.edu/utgsbs_dissertations/995"],"dc:subject":["CRISPR","Functional genomics","Pooled library screening","CRISPR/Cas12a","Biotechnology","Genomics","Medicine and Health Sciences"],"dc:title":["Multiplexed Crispr Libraries For Cancer Functional Genomics"],"thesis:degree_level":["Dissertation (PhD)"],"thesis:degree_name":["Doctor of Philosophy (PhD)"]},"updated_at":"2026-07-24T05:48:47Z"}