羊的肌肉抑制素基因通过CRISPR敲除.pptx

Academic DiscussionCRISPR/Cas9和显微注射技术敲除羊的肌肉抑制素基因Reporter：杨鑫Mentor：王林杰Time：11.09THE MAIN CONTENTS0503ENDDiscussionResultsIntroductionConclusionMaterials/MethodsCONTENT1artPIntroduction01IntroductionMAJOR ADVANCE010203Gene edition has experienced a major advance in the last few years due to the incorporation of novel technologies of direct microinjection into zygotes of gene-specific nucleases such as zinc finger nucleases (ZFN) , TALENs.ProtectionCRISPR/Cas9 Microinjection into ZygotesCRISPR/Cas9 has been rapidly tested in several species to date and promises to revolutionize the field of genome editing in general and in particular transgenic vegetal and animal fields.MyostatinMyostatin is a member of the transforming growth factor beta (TGF-β) superfamily, involved in the inhibition of muscle differentiation and growth.ProductionDiseasesThe objective of this study was to demonstrate the efficiency of the CRISPR/Cas9 gene editing technology coupled to zygote microinjection in the generation of MSTN KO sheep.We show that this approach induces mutations with a high efficiency (45.5%), resulting in live off-spring carrying single-gene mono and biallelic mutations.Introduction2artPMaterials and Methods02Materials and MethodsMethod3artPResults03ResultsResults03ResultsA) Exon 1 of the ovine MSTN gene. The sequence recognized by sgRNA is in italic and bold, from nt 110–130, the PAM sequence TGG is underlined and the rest of exonic sequences in smaller font.B)L: 1Kb DNA. NT:non-transduced cells, T: transfected cells, The experiment is representative of two replicates performed with similar results.Fig 1. CRISPR-Cas9 genome editing activity in transfected ovine cells. Results03ResultsTable 1. Cleavage rate on Day 2 and development rate on Day 6 of embryos that were microinjected into cytoplasm with CRISPR/Cas9 RNA system,buffer injection solution, or non-injected embryos (Control group).Results03ResultsTable 2. Efficiency obtained with CRISPR/Cas9 system injected into cytopl