| Fucosyltransferases (FUTs) are the key enzymes for fucosylatedoligosaccharides synthesis. It has known that stage specific expression ofLey,sLex and Lex oligosaccharides has an important roles in embryodevelopment and implantation. Thus, the studies of effect ofoligosaccharides may provide a new idea and approach to controlimplantation process. A proper and high sensitivity detecting method is important toquantify the gene expression. After comparing quantitative assay amongagarose gel electrophoresis, microfluidic chip and real-time PCR, real-timePCR quantitative was used to analyze FUT2 gene expression in embryosand endometrium and FUTs(FUT1, FUT4, FUT7, FUT8 and FUT9) geneexpression were also studied. Leukaemia inhibitory factor (LIF), which is a secretive glycoproteinwith various biological functions, is a indispensable cytokine duringimplantation stage of mouse, and some implantation-related factors can beregulated by LIF, such as matrix metalloproteinases (MMPs), epidermalgrowth factor (EGF) et.al. LIF was involved in regulation net, but themechanism of the roles of LIF in controlling the expression offucosyltransferase is not clear. In this thesis, the effect of LIF on theexpression of FUT7 was analyzed. The major methods and conclusions in this dissertation aresummarized as the follows: 1. FUT2 and GAPDH gene expression of nonpregnancy and pregnantD4 uterus of mouse were quantified by agarose gel electrophoresis,microfluidic chip after PCR amplification, and the genes also analyzed byreal-time PCR. The sensitivity and detecting limit of these methods wereanalyzed. The results showed that after 25 cycles of PCR amplification, theproduct of FUT2 gene was detected by agarose gel electrophoresis andnone of the product at 20 cycles, whereas the product at 20 cycles was evendetected by microfluidic chip after being diluted by five folds, and thesensitivity of microfluidic chip was 103 times higher than that of theagarose gel electrophoresis. Real-time PCR was a quantitative method withhigh sensitivity and specificity. Gene expression of single embryo could bedetected at 18.88 cycles by real-time PCR , supervising the process at anytime. Real-time PCR was the best in repetition and stability among others. 2. Using real-time PCR to quantify FUT2 gene expression of singleembryos in different developing stage and endometrium of pregnant mouse,the method of quantitative analysis of gene expression of single embryowas improved. The results showed that real-time is a useful quantitativetechnique and it is so sensitive that gene expression of 2-cell stage could bedetected; FUT2 gene expressed on both endometrium and embryo, but thecopy numbers of gene were decreased from about 1011to 1010 or 108 to 107on pregnant D4. The results indicated that FUT2 gene perhaps related withembryo maturation, and construction of receptive endometrium, the variousof gene expression provided experimental evidence for functional study. 3. Using real-time PCR to quantify gene expression of FUT genefamily(FUT1, FUT4, FUT7, FUT8,FUT9)of single embryo ofdifferent developing stage. The results suggested that FUT1,FUT4 andFUT7 gene expression were increased in the course of embryodevelopment, and reach to its peak value at morula stage, expression ofFUT8 gene was stable, and FUT9 gene was expressed in abundance at8-cell, then decreased at morula and blastocyst. The expressions and thetrends of change of FUT gene family were as same as that of the relatedoligosaccharides but were different at developing stage, thus cytokineaffected synthesization and expression of oligosaccharides to controldevelopment of embryo by regulating FUTs gene expression. 4. The function of LIF on FUT7 and sLeX expression of uterusepithelial cell and embryo were studied by PCR, indirectimmunofluorescence and Dot-blot analysis. Embryo and cell werecultured in vitro with medium containing LIF (0.1,1,10,100ng/ml) orLIF-Ab (3μg/ml). The results indicated that LIF could stimulate theexpression of FUT7 on cell and embryo. Effect was obvious when theconcentration of...
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