Porcine VHL gene cloning and construction of VHL knockdown cloned embryos / 生物工程学报

ABSTRACT

Von Hippel-Lindau (VHL) disease is an autosomal dominant disorder and its clinical manifestation including haemangioblastomas of the central nervous system, renal cell carcinoma, haeochromocytomas, and pancreatic cyst. The deletion, mutation and promoter methylation of VHL gene can cause VHL disease. Swine is considered as an ideal model for human disease because of its physiological and anatomical similarity to human. We cloned pig VHL gene that is 2 725 bp in length. VHL highly expressed in adrenal gland, liver, pancreas, heart and testis. We designed 5 shRNAs and screened the most effective interference RNA fragment with a knockdown efficiency of 72%. Porcine embryonic fibroblasts stably transfected with pGenesil-shRNA vector were used as donor cells for nuclear transfer and there was no significant difference of embryo development compared with the control group. Moreover, VHL was efficiently knocked-down with efficiency of 71% in porcine cloned blastocyst, these results lay a solid foundation for constructing the VHL knock-down model of pig.