The Expression Study Of Biological Fusion Protein CTB-LK In Sunflower

Thromboembolic disease is a common cardio-cerebral vascular disease, and it does serious harm to human's life and health. Lumbrokinase has a significant fibrimolytic function in vitro and in vivo. It can dissolve thrombus, reduce relative blood viscosity, inhibite platelet aggregation, degrade the fibrinogen and improve microcirculation, and it shows a good therapeutic effect on the cardio-cerebral vascular disease in clinical.In this study, we used plant bioreactor system--sunflower to express biological fusion protein CTB-LK. Firstly, the two important genes ctb and lk were linked,and subsequently a plant binary expression vector pX6-ctb-lk was successfully constructed, and then transferred into Agrobacterium tumefaciens strain C58. Secondly, sunflower was transformed by C58(pX6-ctb-lk) through agrobacterium-mediated transformation, and 4 kanamycin-resistant plants were regenerated,and some seeds were harvested from 2 of them. We extracted genomic DNA by CTAB method from the leaves of sunflower plant for PCR analysis, the result showed three sunflower plants were positive and it preliminarily showed that the ctb-lk gene had been introduced into sunflower cells. We extracted genomic RNA from the leaves of sunflower plant for RT-PCR analysis and acquired a DNA fragment of correct size. It was confirmed that the ctb-lk gene had integrated into the genome of the transgenic sunflower plants and realized transcription.This is the first report on using sunflower to express to express lumbrokinase fusion protein. Further work such as purification of target protein, bioactivity analysis and animal test needs to be carried out. This study provides technique and theoretical base for using plant system to express foreign proteins and also lays a solid foundation for using transgenic sunflower to express pharmaceutical proteins.