Optimization of Electrotransfection Conditions of EGFP Reporter Gene PiggyBac Vector in Triple Negative Breast Cancer MDA-MB-231 Cell Line
10.3969/j.issn.1671-7414.2025.03.035
- VernacularTitle:EGFP报告基因PiggyBac载体在三阴性乳腺癌MDA-MB-231细胞系电转染条件优化
- Author:
Peipei WANG
1
;
Xiaotian ZHANG
;
Zhiming LING
;
Wenjuan WANG
;
Xiuying LIU
;
Jianxun WANG
Author Information
1. 北京中医药大学生命科学学院,北京 102488;中国医学科学院肿瘤医院山西医院,太原 030000
- Publication Type:Journal Article
- Keywords:
triple-negative breast cancer;
electrotransfection;
MDA-MB-231;
PiggyBac transposon
- From:
Journal of Modern Laboratory Medicine
2025;40(3):189-194
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the optimal conditions for stable transfer of reporter gene containing enhanced green fluorescent protein(EGFP)into human breast cancer cells MDA-MB-231 cells under different electrotransfection conditions.Methods A PiggyBac(PB)transposon system containing EGFP reporter gene was constructed,and transfection conditions such as waveform,voltage,shock time,shock times,plasmid concentration,cell density,transposon to transposase ratio were controlled during electrotransfection of MDA-MB-231 cells.The transfection efficiency was detected by flow cytometry fluorescein isothiocyanate(FITC)channel.The expression of fluorescent protein was observed under laser scanning confocal microscope,the electrotransfection efficiency and expression of EGFP reporter gene were verified,and the electrotransfer efficiency of MDA-MB-231 cells was analyzed under different electrotransfection conditions.Results The optimal transfection conditions of MDA-MB-231 cells were as follows:voltage 280V,exponential wave,electric shock once,concentration of transposon plasmid about 1 000 ng/μl,mass ratio of transposon to transpotase 1:1,number of cells 2×106,electrotransfection rate up to 60.23%±5.63%.The state of MDA-MB-231 cells was good.Conclusion The electrotransfection conditions of MDA-MB-231 cells were successfully optimized to achieve stable and efficient transfection,which provided efficient electrotransfer experimental parameters for the relevant basic research of MDA-MB-231 cell line.
