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Nane Tar More Ko Mor4/21/2021
To install p recise p oint mutation s and to delete or i ntroduc e desired DNA sequences, we highly rely on HD R-mediated precise ge nome editing.
Nane Tar More Ko Mor Download File PDFDownload file PDF Download file PDF Read file Download citation Copy link Link copied Read file Download citation Copy link Link copied Abstract The CRISPRCas9-mediated base editing technology can efficiently generate point mutations in the genome without introducing double-strand break (DSB) or supplying a DNA donor template for homology-dependent repair (HDR).In this study, adenine base editors (ABEs) were used for rapid generation of precise point mutations in two distinct genes, OsWsl5, and OsZebra3 (Z3), in rice protoplasts and regenerated plants.The precisely engineered point mutations were stably inherited to subsequent generations.
Nane Tar More Ko Mor Install P ReciseThese single nucleotide alterations resulted in single amino acid changes and associated wsl5 and z3 phenotypes as evidenced by white stripe leaf and light greendark green leaf pattern, respectively. Through selfing and segregation, transgene-free, base edited wsl5 and z3 mutants were readily obtained in a short period of time. We noticed a novel mutation (V540A) in Z3 locus could mimic the phenotype of Z3 mutation (S542P). Furthermore, we observed unexpected non- AG or TC mutations in the ABE editing window in a few of the edited plants. The ABE vectors and the method from in this study could be used to simultaneously generate point mutations in multiple genes in a single transformation and serve as a useful base editing tool for crop improvement as well as basic studies in plant biology. Nane Tar More Ko Mor Pdf Content AvailableDiscover the worlds research 17 million members 135 million publications 700k research projects Join for free Public Full-texts 2 5dea6cf44585159aa4 6881d9.pdf Content available from CC BY-NC-ND 4.0: 5dea6cf44585159aa46881d9.pdf Simultaneous and precise generation of Zebra3 and Wsl5 mutations in rice using CRISPRCas9-mediated adenine base editors.pdf Content uploaded by Kutubuddin Molla Author content All content in this area was uploaded by Kutubuddin Molla on Dec 06, 2019 Content may be subject to copyright. Simultaneous and precise generation of Zebra3 and Wsl5 mutations in rice using CRISPRCas9-mediated adenine base e ditors.pdf Content available from CC BY-NC-ND 4.0: 5dea6cf44585159aa46881d9.pdf Simultaneous and precise generation of Zebra3 and Wsl5 mutations in rice using CRISPRCas9-mediated adenine base editors.pdf Available via license: CC BY-NC-ND 4.0 Content may be subject to copyright. Molla 1,2 Justin Shih 1 and Yinong Yang 1 1 Department of Plant Patholog y and Environm ental Microbi ology, and th e Huck Institu te of the Life Scien ces, Th e Pennsylvan ia Stat e Universi ty, Univers ity Park, PA 16802, USA. ICAR-National Rice Res earch Institut e, Cuttack-753006, India Emails: KA Molla, kutubudd in.mollaic ar.gov.in J Shih, jws52ps u.edu Y Yang, yuy3 p su. CC-BY-NC-ND 4.0 International license It is made available under a (which was not peer-reviewed) is the authorfunder, who has granted bioRxiv a license to display the preprint in perpetuity. I n this study, adenine b ase editors (ABEs) w ere used fo r rapid ge neration of pr ecise poi nt mutatio ns in two di stinct gene s, OsWsl5, and OsZe bra3 (Z3), in rice protoplast s and regen erated p lants. Th e precis ely engineered point mutations w ere stabl y inherited to subse quent gener ations. Through s elfing and segregation, transgene-free, b ase edited wsl5 and z3 mutants w ere readily obtained i n a short period of time. We no ticed a nove l mutation ( V540A) in Z3 locus could mimic the phenotype of Z3 mutation ( S542P). F urthermore, we obser ved unexpecte d non- AG or TC mutatio ns in the ABE editing window in a few of the edited plan ts. Key Words: Adenine bas e editor, CRISPRCas9, g enome edit ing, plant base editing, precise point muta tion, transg ene-free rice. The b ase editing approach allows rap id generat ion of tran sgene-free ri ce mut ants with exp ected phenotypic ch anges.. The discovery of th e CRISPRCas9 s ystem and its repurposing for gen ome editin g revolutionized basic biol ogical rese arch and practic al applicatio ns in medic ine and agric ulture (Do udna and Charpenti er, 2014). In th e CRISPRCas9 -mediated genome editin g process, a single guide R NA (sgRNA ) can direct Cas9 to create a double strand break (DSB) at a target l ocus with high precisi on. Higher euka ryotes includ ing plant c ell repairs th e DSB through eith er non- homologous end joining (NHEJ ) or homology direct ed repair (HDR) pat hways (Molla an d Yang, 2019 a ). Higher plants predominan tly use the err or prone NH EJ which c reates ran dom insertio ndeletion ( indel) caus ing frame shift mutatio n and ultim ately ge ne knock out (Huang an d Puchta, 20 19). NHEJ-media ted gene knock -out has limi ted applicat ion in crop im provement s ince it cannot cre ate precise indels or specific po int mutatio ns for soph isticated cr op genome engineerin g.
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