cell culture 07
TRANSCRIPT
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Advanced Aminal Cell Culture
2013 2nd Semester
Department of Animal Science
Chungbuk National University
7th Lecture
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Syllabus
Date TopicsSeptember 5, 2013 Introduction : What is Cell Culture?
September 12, 2013 Cell Culture As Model System For Research
September 26, 2013 Cell Culture For Antibody / Protein Production
October 17, 2013 Protein Production/Purification
October 31, 2013 Stem Cell INovember 14, 2013 Stem Cell IINovember 28, 2013 TG/KO Animals December 5, 2013 Genome Engineering/NGS
December 12, 2013 Final Exam
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DateSeptember 26, 2013 Cell Culture For
Antibody / Protein Production
조유미 , Madhusumida
October 17, 2013 Protein Production/Purification 이미진 , 정용호
October 31, 2013 이영 , 윤준호November 14, 2013 Stem Cell I Jia Jia Lin, 염동현
November 28, 2013 Stem Cell II Zhao MingHui,권정우December 5, 2013 Transgenic Animals Lin Zili, 이상배December 12, 2013 Genome
Engineering/NGS 조유진 , 김상욱
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Knockout/Knock In/Transgenic Animals
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Why we care about that?
- Knockout/Knock-In animal (mouse) : Disease Model For Medical Research
- Immune-deficient Pig : Xenotransplantation?
- Transgenic Animals : Animal Bioreactor?
Production of useful protein in Milk or Egg?
- Domestic Animal Breeding?
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Knockout Mouse
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Knockdown vs KnockoutGene Knockdown
- Expression level of mRNA was decreased by RNAi (siRNA/shRNA/dsRNA)- Gene itself are still intact- SilencingEfficiency is not ~100%- Sometime residual activities are sufficient for phenotypes- Relatively easy and fast to perform- You can do in embryonic lethal gene
Gene Knockout
- Gene locus in chromosome is permanently removed/changed
- Complete loss of genotype- Difficulty & Time consuming- Embryonic lethal gene : Conditional Knockout
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Knockout mouse from Embryonic Stem Cell
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Homologous Recombinantion
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BAC recombineering
BAC : Bacterial Artificial Chromosome
Plasmid vector can have ~150kb of insertUsed for the Original Human Genome ProjectsIdeal for modification of Large fragments for the homologus recombinantion
Most of
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Conditional Knockout
- Cre recombinase
- Why we need to make conditional knockout mouse ?
Embryonic Lethal
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Oocyte Specific Knockout
- Cre recombinase are expressed in oocyte by ZP3 promoter- Desired locus has modified to have LoxP
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Knock inTransgenic vs Knock-In?
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Transgenic Animals
- Embryonic Stem Cell
- Somatic Nuclear Transfer
- Viral Vector
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Novel strategy for generation of knockout/knock-in/transgenic
- Zinc-Finger Nuclease- TALEN- CRISPER
Genome Engineering?
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Zinc-Finger Nuclease
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TALEN
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CRISPR
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One-step generation of Knockout Animal using TALEN