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[Dual gene amplification and selection system with dihydrofolate reductase and glutamine synthetase genes effectively
Zhiyun Wang1, Bo Wei, Shufang Tian
1Xi'an Jiaotong University, Xi'an 710061, China.
Background:
To study the effect of gene amplification and selection system with DHFR plus GS and DHFR or GS gene on the foreign gene expression.
Methods:
Using the N-terminal truncated hTPO(T184) gene as target gene, two plasmidsre were constructed: pDC- T184 and pGC-T184 where DHFR and GS gene were used respectively as the selective amplification marker. They were cotransfected into CHO dhfr cells to establish dual gene amplification and selection system of DHFR plus GS gen and respectively transfected to establish single gene amplification and selection system of DHFR or GS gene. Three selective methods in dual selective system to compare expression efficiency of hTPO were designed: the first method (DG) was to use drug pressure of MTX, then use MSX; the second method (GD) was reversed; the third method was simultaneously to use MTX and MSX as drug pressure.
Results:
DHFR+GS dual system had not only higher gene amplification efficiency but also higher level expression. There was no distinct affect in different method of drug pressure.
Conclusions:
MTX plus MSX dual drug pressure in dual selection system was an efficient and simple method to increase the expression of foreign gene in mammalian cells.