本文選題：免疫性血小板減少癥　切入點(diǎn)：凝血因子V　出處：《廣西醫(yī)科大學(xué)》2012年碩士論文

【摘要】：目的探討兒童免疫性血小板減少癥(ITP)發(fā)病及病情與血小板膜糖蛋白(GP1BA)基因、FV因子第10外顯子(FV10)、VWF基因第28外顯子(VWF28)變異相關(guān)性,以求對明確兒童ITP發(fā)病與此基因變異有無的遺傳相關(guān)性,探索ITP發(fā)病的遺傳機(jī)制。 方法對51例臨床診斷為兒童ITP的患兒和28例非ITP患兒的外周血進(jìn)行全基因組DNA提取,參照CCDS數(shù)據(jù)庫人類基因序列設(shè)計PCR引物,以基因組DNA為模板對GP1BA、FV10、VWF28基因進(jìn)行PCR擴(kuò)增、凝膠電泳、基因測序,依據(jù)NCBI等數(shù)據(jù)庫信息整理兩組數(shù)據(jù)進(jìn)行統(tǒng)計學(xué)分析。 結(jié)果(1)GP1BA基因存在T161M,(24.0%,5.97%,P=0.011)、W614R氨基酸變異點(diǎn),檢測樣本基因存在大段缺失現(xiàn)象,發(fā)現(xiàn)GP1BA基因有重復(fù)串聯(lián)現(xiàn)象。(2)成功擴(kuò)增實(shí)驗(yàn)組及對照組所有患兒FV10基因；與數(shù)據(jù)庫FV基因及氨基酸序列比結(jié)果顯示ITP組和對照組均存在氨基酸R513K變異(94.1%、100%,P=0.191),雜合子率為87.5%、78.6%(P=0.303)；兩組氨基酸均存在Q534R變異,且變異率為100%；未發(fā)現(xiàn)FV氨基酸506位的FVLeiden突變(核苷酸G1691A變異\氨基酸A506G變異)；(3)成功擴(kuò)增90%以上樣本VWF28基因全序列,發(fā)現(xiàn)V1229I、V1229G雜合、N1231T雜合、G1253S雜合、W1313R雜合、T1381A變異(約78%)、D1472H雜合、E1376G雜合、V1565L氨基酸變異共9種氨基酸變異。GCC---TTG脫氧核糖核酸具有連鎖遺傳特性,V1565L的變異不論是純合狀態(tài)還是雜合狀態(tài),100%都會與A1555A(GCC、GAA) (?)勺雜合堿基無義突變連鎖出現(xiàn)； 結(jié)論(1)GP1AB基因上的T161M的變異與兒童ITP的發(fā)生相關(guān)。(2)GP1BA的W614R變異,可能與患兒ITP的發(fā)生有關(guān)。(3)FV發(fā)生的R513K、Q534R氨基酸變異與兒童ITP發(fā)生無相關(guān)性。(4)ITP發(fā)生與FV Lieden突變無相關(guān)性。(5)兒童ITP的發(fā)生可能與VWF28基因變異有關(guān)；VWF28上V1229I,V1229G,N1231T,G1253S,W1313R,E1376G等變異,可能與患兒ITP的發(fā)生有關(guān)。T1381A、D1472H變異、E1376G變異與兒童ITP發(fā)生無關(guān)。(6)廣西地區(qū)人口VWF28基因存在GCC---TTG脫氧核糖核酸的連鎖遺傳；(7)部分兒童ITP發(fā)生與基因變異有關(guān)；尚不能確定兒童ITP與FV、VWF、GPIBA全基因變異的相互作用。
[Abstract]:Objective to investigate the relationship between the pathogenesis and condition of ITPs in children with immune thrombocytopenia and the variation of VWF28 in exon 10 of FV10 exon 10 of platelet membrane glycoprotein (GP1BA) gene.In order to find out the genetic correlation between ITP and the gene variation in children, and to explore the genetic mechanism of ITP.Methods Genomic DNA was extracted from peripheral blood of 51 children with clinically diagnosed ITP and 28 children without ITP. PCR primers were designed according to the human gene sequence of CCDS database. Genomic DNA was used as template for PCR amplification of GP1 BAFV10VWF28 gene.Gel electrophoresis, gene sequencing, NCBI and other database information collate the two groups of data for statistical analysis.Results the amino acid mutation of W614R was detected at 5.97 and 5.97. The FV10 gene of all the children in the experimental group and the control group was amplified successfully. The results showed that there was a large deletion in the sample gene and the GP1BA gene was duplicated and connected with the other two genes in the experiment group and in the control group, and the FV10 gene of all the children in the experimental group and the control group was amplified successfully.Compared with FV gene and amino acid sequence in database, the amino acid R513K mutation was found in ITP group and control group. The amino acid R513K mutation was 94.1and 0.191%, the heterozygote rate was 87.5% and 78.6% (P < 0.05), Q534R mutation was found in both groups.There was no FVLeiden mutation (nucleotide G1691A mutation / amino acid A506G mutation) at position 506 of FV amino acid to amplify the complete sequence of VWF28 gene in more than 90% of the samples.鍙戠幇V1229I,V1229G鏉傚悎,N1231T鏉傚悎,G1253S鏉傚悎,W1313R鏉傚悎,T1381A鍙樺紓(綰，

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