The Application Of The Quantified Trans-placental RNAi Technology In Mouse Model’s Establishment

ObjectiveTo assess a efficient technology to establish knock-down mouse bytrans-placental RNAi-BMP-4(Bone morphogenetic protein) to mouseembryos. In2006, O’Shea found that the pregnant mice interferred withsingle-gene plasmid in the early phase of mouse embryos could cause thedevelopmental disorder of tissue and organ in the offspring mice andproduce deformity mice by knocking down the target genes viatrans-placental effect. With the support of Natural Science Foundation ofChina (NSCF) basis by the preliminary work, our research group took afive-year study to master the trans-placental RNA interference technology,firstly set up and optimized the platform of the relevant technologies, andusing the technology to build some congenital disease model animals.MethodsAfter established BMP-4down-regulatory plasmid, the Ringer’s,pSES and pSES-SiBMP-4are delivered to pregnant mouse via tail vein.Observed the growth and development of embryos after48h, then usingRT-PCR and Western blotting analysis it. Observed the newborn mouse, then diagnosis disease using matching method.ResultsCompare to the Ringer’s group and pSES group, there was areduction in BMP-4mRNA and BMP-4protein in experimental group (P＜0.05). There is no significant malformation between Ringer’s group andcontrol group. In pSES-SiBMP-4group mouse embryos, head,gastrointestinal tract and backbone seem abnormal, also epilepsy, polypsand Hirschsprung are found in offsprings.ConclusionsIt is feasible to deliver plasmid to early staged mouse embryos that setup and optimized the platform of the relevant technologies. It can be usedto evaluate gene function and create the downregulated model animal.(1) RNAi plasmid could crossed the early stage placental anddown-regulated the offspring gene expression. The effects is related to theinterference plasmid injection time window, plasmid quantity, plasmidconcentration, injection speed.(2) BMP-4gene is a key role to the growthand development of the mouse in nervous system (CNS/ENS), the skeletalsystem, circulatory system, the digestive system and skin.(3) qTP-RNAican be quantitatively reduced offspring’S BMP-4gene. Regulation theinjection conditions can building different congenital disease’s animalmodels--Congenital epilepsy, colon polyps and Hirschsprung Model.