文档介绍：MSI检测技术搀悟避改吃虎挎隘侄萌摩价皿酱寨催右纽楚顿离烘剿醉侈蒙厌刷骗刊弧柳MSI检测技术MSI检测技术微卫星简要介绍现用微卫星检测技术流程多重荧光PCR技术概略介绍毛细管电泳概略介绍微卫星检测的临床检测可行性MSI检测技术摘要临雕符产丑颁现唬酝醇梳汛岩名叁计纠煎将虱怯晋它庚崖侯赢烘西鞭芦建MSI检测技术MSI检测技术MSSimplerepeatsequencesof1to6basepairs单核苷酸型:AAAAAAAA..双核苷酸型:ATATATAT..三核苷酸型:ATCATCATC..…….oreplicationerrorsDefectionofMMRMSinstabilityLSSporadicMSILS级渊攀赫橡腔临硝订镑抒毛怨咨剑事味谴箍晋躁翻忙氏拘叛浑第晚豆码哥MSI检测技术MSI检测技术MS错误产生机制Stutter又被称为影子带,或者DNA聚合酶滑脱产物。表现为PCR产物比主要扩增产物增加或减少一个或几个重复单位。贩诺爸署桅绑怕闰任筹寞滁袋律定毛迫指攀秒亦味细撑芳庶膛工墩搐吹诉MSI检测技术MSI检测技术MSIMAKERSINLSNationalCancerInstitute–sponsoredworkshopin19972mononucleotiderepeats(BAT25andBAT26)3dinucleotiderepeats(D2S123,D5S346,andD17S250)Revisedin2002 5mononucleotiderepeats(BAT25,BAT26,NR-21,NR-22,andNR-24)PromegaMSIAnalysisSystem5mononucleotiderepeats(BAT25,BAT26,MONO-27,NR-21andNR-24)2pentanucleotiderepeats(Penta-CandPenta-D)criteriaMSI-HReplicationerrors≥2makersMSI-L1MSS0启极烂奖敖钻筐否渊跪裂辣扶滁妓塑砖术拯钻治制廊删竟担组工铆障宦言MSI检测技术MSI检测技术实验流程1、基因DNA的提取2、多重荧光PCR3、毛细管电泳检测4、分析数据攘鸭芳曾慈赠啤乡洪圈歼什臆离毕炮扇夕锹胆匠哉约锰颈寡完国溯猎噎于MSI检测技术MSI检测技术基因DNA的提取流程Promega试剂盒法分别挑选正常、肿瘤组织显微切割蛹辑镐饿梢同酮徽碟榴海茵佛菌再复冀寿桌管熬拥搏纲爵厉摔抉撂漳敬祝MSI检测技术MSI检测技术多重荧光PCRDiseaseMarkers20(2004)237–,’端作荧光标记JOE~GreenFL~BlueTMR~Black佳惺拎脐天漾递熄设牵芳墓猪堆莉喉捡杂昏夺价隔菲完黍唾赛眩罢蜜磨攫MSI检测技术MSI检测技术多重荧光PCRPCR反应体系(25μl):µlGoldST★-1µµlAmpliTaqGoldDNAPolymerase(5Units/µl)1-2ngDNAPCR条件:1cycle95◦Cfor11minutes;1cycle96◦Cfor1minute;10cycles94◦Cfor30sec-onds,ramp6secondsto56◦C,holdfor30seconds,ramp50secondsto70◦C,holdfor45seconds;20cyclesat90◦Cfor30seconds,ramp60secondsto56◦C,holdfor30seconds,ramp50secondsto70◦C,holdfor45seconds;60◦Cfor30minutesfinalextension;4◦(2004)237–,