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miR-30a Can Effect the Expression of Liver Specific Gene on HepG2Cells

Author: WangLiuYi
Tutor: FengYiChao; HanYing
School: Yan'an University
Course: Internal Medicine
Keywords: miR-30a Albumin HepG2cell Interference sequence lentiviral vector mimics
CLC: R730.2
Type: Master's thesis
Year: 2013
Downloads: 3
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Abstract


【Background and Purpose】MicroRNA (miRNA) has been found in eukaryotes and it has a class of about19to24nucleotides in non-coding single-stranded RNA molecules, a large number of miRNAs indifferent species have been reported one after another. miRNA can regulate thedifferentiation and development process of many cells. Transcription factor can not onlyregulate the development of cells, it can also participate in the regulation of theexpression of albumin which is one of the liver-specific genes. Not only the transcriptionfactor has this function,the miRNA also involved in the regulation of liver development.Our group already established a differentiation model of human umbilical cordmesenchymal stem cells differentiate into the hepatocyte.We choose the cell samples atdifferent time points in the differentiation process, combined with gene chip technology,and qRT-PCR technology to filter out a group of miRNA expression profiling in hepaticspecifi differentiation.Among this MiR-30a is over-expression. Combining this resultwith review of the literature, we choose miR-30a,which is relate to the selection anddevelopment of liver cell and bile duct cell development, and we choose a humanhepatoma cell line HepG2cell lines to investigate the expression of miR-30a in this cellline,and wheth miR-30a can affect the expression of liver-specific genes in human serum albumin.【Method】Choosing HepG2as a seed cells,and construct lentiviral vector of miR-30a interferencesequence to infect this cell to detect infected with the virus by real-time quantitativePCR.We want to see wheth miR-30a can affect the expression of liver-specific genesALB in HepG2cells.And we use miR-30a mimics to transfected with stable transfectionof miR-30a virus in HepG2cells,we want to explore if we restore the expression ofmiR-30a in HepG2cells, miR-30a can also continue to affect the expression of ALB. 【Result】1、 We has infected the interference sequence miR-30a lentivirus vector into HepG2cells,and we also established miR-30a cell whic can stable transfection in HepG2cellsand successfully.2、 When we transfected the mimics into the HepG2cells which stable transfectionwhith miR-30a, this can elevate the expression levels of miR-30a in HepG2cells(p <0.05).3、 Low expression of miR-30a can inhibit the expression of liver-specific gene ALB inHepG2cells(p <0.05).4、 If we restore the expression levels of miR-30a in HepG2cells, the expression ofliver-specific gene ALB is also rising(p <0.05).【Conclusion】In this study,we follow the preliminary findings, choose the miR-30a which relatewhith the development of hepatocyte and bile duct cells.we choose HepG2cells for thepurpose of our seed cell.Constructed a ltiviral vectors of miR-30a and miRNA mimics,using qRT-PCR technology from the mRNA level, we find low expression of miR-30acan inhibit the expression of liver-specific gene ALB. Taking the cell which has stabletransfected of miR-30a virus, transfected with miR-30a mimics into this cell sample,wefound that when the recovery miR-30a expression levels, the expression of ALB also willincrease,.This tould that miR-30a can maintaine the expression of ALB in HepG2cellsUsing western blot can also confirme this. The results of this study provide a basis for theregulation of liver-specific gene expression and its mechanism by miRNA.

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