SHERP

Comparison of Various Transfection Methods in Human and Bovine Cultured Cells
인간 및 소 배양 세포에서 다양한 형질주입 방법의 비교

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Authors
김용훈
Advisor
노상호
Major
자연과학대학 협동과정 유전공학전공
Issue Date
2014
Publisher
서울대학교 대학원
Keywords
transfectionmagnetofectionnon-liposomal transfectionelectroporation형질주입자기주입형질주입전기천공
Description
학위논문 (석사)-- 서울대학교 대학원 : 협동과정 유전공학전공, 2014. 8. 노상호.
Abstract
형질주입은 유전자전달방법으로서 세포의 성상을 조절할 때 널리 이용되는데 ‘Yamanaka 인자’ 라고 하는 세포 재프로그래밍 인자를 형질주입하여 유도만능줄기세포를 생산하는 것을 예로 들 수 있다. 하지만 형질주입 효율은 일반적으로 낮아 개선이 필요하고 더 우수한 형질도입 방법을 도입할 필요가 있다. 본 연구에서는 non-liposomal 형질주입법 (NLT), 자기주입법 (magnetofection) 및 전기천공법 (electroporation)을 이용하여 인간치수세포 및 소태아섬유모세포에 green fluorescence protein (GFP)을 전달하여 각 방법의 형질주입 효율 및 생존효율을 분석하였다. 자기주입법으로서 PolyMag 을 사용하였고 NLT 로서 Fugene-HD 및 X-tremeGENE 9 을 사용하였으며 전기천공법으로서 NEPATM 및 NeonTM 전기천공기를 사용하였다. NeonTM 전기천공법은 NLT, 자기주입법 및 NEPA TM 전기천공법와 비교하였을 때 인간치수세포에서는 33% 소 태아 섬유아세포에서는 50%로 가장 높은 형질주입 효율을 나타내었다. 본 연구결과로 인간치수세포와 소태아유아세포에서 Neon TM 전기천공법이 가장 효율적인 형질주입법이라는 것이 확인되었다.
Transfection is a gene delivery tool and widely used to manipulate cellular property such as induced pluripotent stem cell (iPSC) generation by reprogramming factors so called ‘Yamanaka factors’. However, transfection efficiency still needs to be improved and more efficient method is required. In the present study, three transfection protocols including non-liposomal transfection (NLT), magnetofection and electroporation were compared by analyzing their transfection efficiencies and cell viabilities using human dental pulp cells (hDPC) and bovine fetal fibroblasts (bFF) as cell sources. Enhanced green fluorescent protein gene was used as delivery indicator. For magnetofection, Polymag reagent was administrated, and for NLT, FuGENE-HD and X-treme GENE 9 DNA transfection reagent were used. For electroporation, two electroporators called NeonTM and NEPA21TM were tested. NeonTM electroporation showed highest transfection efficiency when compared with NLT, magnetofection, and NEPA21TM electroporation, exhibiting approximately 33% of transfection efficiency in hDPC and 50% in bFF, based on viable cell population in each cell type. These results suggest that transfection by NeonTM electroporation can be used to deliver foreign genes efficiently in human and bovine somatic cells.
Language
English
URI
http://hdl.handle.net/10371/131163
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College of Natural Sciences (자연과학대학)Program in Genetic Engineering (협동과정-유전공학전공)Theses (Master's Degree_협동과정-유전공학전공)
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