S-Space College of Agriculture and Life Sciences (농업생명과학대학) Dept. of Biosystems and Biomaterials Science and Engineering (바이오시스템·소재학부) Theses (Master's Degree_바이오시스템·소재학부)
Development of a Bioelectrospray System Enhanced Gene Transfection into Mammalian Cells
세포 내 유전자 도입 효율증대를 위한 바이오전기분무 시스템 개발
- LEE MYUNG CHUL
- 농업생명과학대학 바이오시스템·소재학부
- Issue Date
- 서울대학교 대학원
- Transfection; Electrospray; Bio-electrospray(BES); Non-viral vector; Polyethylenimine (PEI)
- 학위논문 (석사)-- 서울대학교 대학원 : 바이오시스템·소재학부(바이오시스템공학전공), 2015. 8. 정종훈.
- To deliver genes into animal cells, viral vectors are generally used because of their high transfection efficiency. However, viral vectors have a risk that it can infect pathogenic genes into the host. Thus, many researchers use non-viral vectors such as synthetic polymers and apply mechanical method. However, their transfection efficiency are lower than that of viral vectors.
In this paper, we tried to combine chemical and mechanical methods for enhancing transfection efficiency. Cationic polymers were selected as chemical method. Then Mechanical method was applied by new way of transfection. The new method is bio-electrospray (BES). The bio-electrospray (BES) is an application of electrospray. This is technique to electrospray living cells on target. We formed a hypothesis that gene transfection efficiency can be increased if the bio-electrospray (BES) method is combined with chemical method (using polymer). After that, as procedures to increase the gene transfection efficiency in animal cells, we tried to combine polyethylenimine (PEI), a cationic polymer (non-viral carrier) with bio-electrospray (BES) system.
At first, we evaluated cell viability after BES. In order to investigate what solvent elevates transfection level, vector-PEI complex and cells were mixed with several solvents [phosphate buffer saline (PBS), deionized water (DW), Dulbeco's Modified Eagle's Medium (DMEM), and culture media (CM)]. Then in order to investigate what voltage elevate transfection level, the mixtures were electrosprayed with various voltages. Their transfection efficiency is examined by turbo green fluorescent protein (tGFP) expression and luciferase assay.
As a result, when PBS as solvent that mixed cells, DNA and PEI was applied, transfection level was much higher than others. Moreover, when 5kV was applied, transfection level is the highest.
From this study, the new way of transfection was presented and it will be applied at transfection of non-adhesion cells like lymphocyte.