摘要
The antisense fragments, which were available in in vitro system, were cloned into the mammalian expression vector pcDNA3, and were transfected into H654 cells, a mammalian cell line stably expressing the thalassaemic (IVS 2 654 C→T) human β globin gene. In these transfected cells, the level of correctly spliced β globin mRNA in total β globin mRNA (β/(β+β *)) was improved from 0.07 (0 d) to 0.22 (3 d), and this effect persisted for up to 15 d post transfection. All the results demonstrated that antisense RNAs were able to be transcribed from the antisense fragment expression vectors stably and effectively suppressed aberrant splicing pattern of the mutated β globin gene (IVS 2 654 C→T) and restored correct splicing pathway. This work provided a novel approach with potential clinical significance to gene therapy of this kind of splicing mutants including β thalassaemia (IVS 2 654 C→T) by antisense RNAs.
The antisense fragments, which were available in in vitro system, were cloned into the mammalian expression vector pcDNA3, and were transfected into H654 cells, a mammalian cell line stably expressing the thalassaemic (IVS 2 654 C→T) human β globin gene. In these transfected cells, the level of correctly spliced β globin mRNA in total β globin mRNA (β/(β+β *)) was improved from 0.07 (0 d) to 0.22 (3 d), and this effect persisted for up to 15 d post transfection. All the results demonstrated that antisense RNAs were able to be transcribed from the antisense fragment expression vectors stably and effectively suppressed aberrant splicing pattern of the mutated β globin gene (IVS 2 654 C→T) and restored correct splicing pathway. This work provided a novel approach with potential clinical significance to gene therapy of this kind of splicing mutants including β thalassaemia (IVS 2 654 C→T) by antisense RNAs.
基金
ProjectsupportedinpartbytheNationalNaturalScienceFoundationofChina (GrantNos .39780 0 1 9
39392 90 3)
theShanghaiLifeSciencesResearchCentre .
