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Simplified homology-assisted CRISPR for gene editing in Drosophila.
Rankin, Anne E; Fox, Elizabeth; Chisholm, Townley; Lantz, Nicole; Rajan, Arjun; Phillips, William; Griffin, Elizabeth; Harper, Jaekeb; Suhr, Christopher; Tan, Max; Wang, Jason; Yang, Alana; Kim, Ella S; Ankrah, Naa Kwama A; Chakraborty, Praachi; Lam, Alistair C K; Laws, Madeleine E; Lee, Jackson; Park, Kyle K; Wesel, Emily; Covert, Peter H; Kockel, Lutz; Park, Sangbin; Kim, Seung K.
Afiliação
  • Rankin AE; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Fox E; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Chisholm T; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Lantz N; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Rajan A; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Phillips W; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Griffin E; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Harper J; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Suhr C; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Tan M; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Wang J; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Yang A; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Kim ES; Phillips Exeter Academy, Exeter, NH 03833, USA.
  • Ankrah NKA; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Chakraborty P; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Lam ACK; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Laws ME; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Lee J; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Park KK; The Lawrenceville School, Lawrenceville, NJ 08648, USA.
  • Wesel E; Stanford University, Stanford, CA 94305, USA.
  • Covert PH; Stanford University, Stanford, CA 94305, USA.
  • Kockel L; Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
  • Park S; Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
  • Kim SK; Department of Developmental Biology, Stanford University School of Medicine, Stanford, CA 94305, USA.
G3 (Bethesda) ; 14(2)2024 Feb 07.
Article em En | MEDLINE | ID: mdl-38058125
In vivo genome editing with clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 generates powerful tools to study gene regulation and function. We revised the homology-assisted CRISPR knock-in method to convert Drosophila GAL4 lines to LexA lines using a new universal knock-in donor strain. A balancer chromosome-linked donor strain with both body color (yellow) and eye red fluorescent protein (RFP) expression markers simplified the identification of LexA knock-in using light or fluorescence microscopy. A second balancer chromosome-linked donor strain readily converted the second chromosome-linked GAL4 lines regardless of target location in the cis-chromosome but showed limited success for the third chromosome-linked GAL4 lines. We observed a consistent and robust expression of the yellow transgene in progeny harboring a LexA knock-in at diverse genomic locations. Unexpectedly, the expression of the 3xP3-RFP transgene in the "dual transgene" cassette was significantly increased compared with that of the original single 3xP3-RFP transgene cassette in all tested genomic locations. Using this improved screening approach, we generated 16 novel LexA lines; tissue expression by the derived LexA and originating GAL4 lines was similar or indistinguishable. In collaboration with 2 secondary school classes, we also established a systematic workflow to generate a collection of LexA lines from frequently used GAL4 lines.
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Texto completo: 1 Base de dados: MEDLINE Assunto principal: Drosophila / Edição de Genes Limite: Animals Idioma: En Ano de publicação: 2024 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Drosophila / Edição de Genes Limite: Animals Idioma: En Ano de publicação: 2024 Tipo de documento: Article