Designation and silencing function of the small interfering RNA of HLA-A2
10.3969/j.issn.2095-4344.2014.29.003
- VernacularTitle:HLA-A2小干扰RNA的设计及沉默效果检测
- Author:
Jiansheng LIU
;
Gang ZHAO
;
Dan PEI
;
Deyu ZHENG
- Publication Type:Journal Article
- Keywords:
HLA antigens;
RNA interference;
gene silencing;
fibroblasts
- From:
Chinese Journal of Tissue Engineering Research
2014;(29):4605-4610
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUND:Human leukocyte antigen (HLA), the major histocompatibility complex of human, plays an important function in the transplant rejection. Decreasing the expression of HLA wil prolong the survival time of transplants.
OBJECTIVE:To design smal interfering RNA (siRNA) of HLA-A2 and to detect the effect of siRNA-HLA-A2 on the expression of HLA-A2.
METHODS:Four kinds of siRNA-HLA-A2 domains were designed, and recombinant lentivirus expression vector were formed. The 293T cells, highly expressing HLA-A2, were infected in vitro. Then the knockout efficacy of four domains was detected to select the highly efficient siRNA-HLA-A2 target sequences. The human embryo lung fibroblasts were cultured in vitro and infected with the lentivirus carrying the target sequence. The infecting efficiency of LV-siRNA-HLA-A2 was observed under the fluorescence microscope and the silence function of this siRNA in human embryo lung fibroblasts was detected by western blot analysis.
RESULTS AND CONCLUSION:According to the mRNA sequence of HLA-A2 in Genbank, three siRNAs were designed and synthesized. In vitro, the over expression of HLA-A2 in 293K cells was successful y silenced. The HLA-A2 expression in human embryo lung fibroblasts was also efficiently silenced after the human embryo lung fibroblasts were infected by the highly efficient siRNA of HLA-A2. The efficacy was up to 80%.
