本文選題：Y-STR + 復(fù)合擴(kuò)增　； 參考：《四川大學(xué)》2003年博士論文

【摘要】： 目的 Y染色體STR(Y-STR)呈單倍型父系遺傳特征，不僅在人類(lèi)學(xué)、古生物學(xué)等方面意義重大，而且在法醫(yī)學(xué)的個(gè)人識(shí)別及親子鑒定中亦具有重要而獨(dú)特的應(yīng)用價(jià)值。本課題旨在通過(guò)研究復(fù)合擴(kuò)增技術(shù)，構(gòu)建四個(gè)Y-STR基因座的復(fù)合擴(kuò)增銀染檢測(cè)體系和復(fù)合擴(kuò)增熒光檢測(cè)體系，并對(duì)其進(jìn)行法醫(yī)學(xué)可行性研究和群體遺傳研究。以期為法醫(yī)Y染色體分型提供新的技術(shù)手段。方法 選定四個(gè)Y染色體STR基因座，應(yīng)用加尾序列引物設(shè)計(jì)策略設(shè)計(jì)的引物，構(gòu)建四個(gè)基因座的Y-STR復(fù)合擴(kuò)增體系，建立銀染檢測(cè)和熒光檢測(cè)方法，依據(jù)DNA分析技術(shù)工作組(TWGDAM)指南進(jìn)行法醫(yī)學(xué)可行性研究和群體遺傳研究。結(jié)果 構(gòu)建了四個(gè)Y-STR基因座：A10(Y-GATA-A10)，DYS434，DYS438，DYS439復(fù)合擴(kuò)增體系(Y-A489-plex)；建立了Y-STR復(fù)合擴(kuò)增銀染檢測(cè)方法和熒光檢測(cè)方法。法醫(yī)學(xué)可行性研究結(jié)果顯示，敏感度檢驗(yàn)最低模板量Y-A489-plex銀染體系為0.5ng，Y-A489-plex熒光體系為0.125ng；分型結(jié)果穩(wěn)定，重復(fù)性好，對(duì)陳舊斑痕具有檢測(cè)能力，能夠?qū)ΤＲ?jiàn)介質(zhì)上的斑痕正確分型， 具有很好組織同一性、種屬特異性，可以耐受女性成分的干擾， 用于混合斑分析獨(dú)具優(yōu)勢(shì);可以用于實(shí)際檢案。在100個(gè)樣本中 檢出37個(gè)單倍型，單倍型變異度為0.9628，標(biāo)準(zhǔn)誤為0.00429。 結(jié)論本研究在國(guó)內(nèi)外第一次將加尾序列引物設(shè)計(jì)策略引入 Y一STR復(fù)合擴(kuò)增體系。證明該引物設(shè)計(jì)策略在Y一STR復(fù)合擴(kuò)增研 究中是切實(shí)可行的。加尾引物復(fù)合擴(kuò)增策略還有很大的潛力，為 大規(guī)模Y一STR復(fù)合擴(kuò)增的研究提供了一個(gè)行之有效的方法。 初步構(gòu)建了四個(gè)Y一STR基因座的Y一A489一plex銀染體系和 Y一A4Sg一plex熒光體系:并依據(jù)DNA分析技術(shù)工作組(TWGDAM) 指南進(jìn)行了應(yīng)用性研究。證明Y一A489一plex，適合于法醫(yī)DNA分 析，有較高的應(yīng)用價(jià)值，，為法醫(yī)Y染色體分型提供了新的技術(shù)手 段。 Y一A489一plex體系采用的是國(guó)產(chǎn)的Taq酶。經(jīng)與美國(guó)生產(chǎn)的 金牌酶比較，無(wú)論是擴(kuò)增效率還是特異性都無(wú)明顯差異，為試劑 國(guó)產(chǎn)化進(jìn)行了有效的探索。
[Abstract]:Objective Y chromosome STR- Y-STR has haplotype paternal genetic characteristics, which not only has great significance in anthropology and paleontology, but also has important and unique application value in forensic personal identification and paternity test. The purpose of this study was to construct a multiplex amplification silver staining detection system and a multiplex amplified fluorescence detection system for four Y-STR loci, and to study the feasibility of forensic medicine and population heredity of the four Y-STR loci. In order to provide a new technique for forensic Y chromosome typing. Methods four STR loci of Y chromosome were selected. The Y-STR multiplex amplification system of the four loci was constructed by using the primers designed by adding tail sequence primers, and the silver staining and fluorescence detection methods were established. Forensic feasibility studies and population genetics studies were conducted in accordance with the TWGDAM guidelines of the working Group on DNA Analysis techniques. Results four Y-STR loci were constructed, and the multiplex amplification system of DYS434 (DYS438) DYS439 was constructed, and the silver staining and fluorescence detection methods of Y-STR multiplex amplification were established. The results of the feasibility study of forensic medicine showed that the minimum template quantity of sensitivity test was 0.5ngA489-plex silver staining system, the fluorescence system of Y-A489-plex fluorescence system was 0.125ng.The typing results were stable, reproducible, and had the ability to detect the old stains. Able to type correctly the stains on common media, with good tissue identity, species specificity, It can tolerate the interference of female component, and can be used for mixed spot analysis. 37 haplotypes were detected in 100 samples, the variation of haplotypes was 0.9628, and the standard error was 0.00429. Conclusion for the first time, the design strategy of tail sequence primer was introduced into Y-STR compound amplification system at home and abroad. It is proved that the primer design strategy is feasible in the study of Y-STR compound amplification. The strategy of adding tail primers has great potential, which provides an effective method for the study of large scale Y STR multiplex amplification. Y-A489 plex silver staining system and Y A4Sg plex fluorescent system were constructed with four Y STR loci. The guide conducts an application study. It is proved that Y A 489 complex is suitable for forensic DNA analysis and has high application value. It provides a new technical hand segment for forensic Y chromosome typing. Y-A489- plex system Domestic Taq enzyme was used. Compared with the gold enzyme produced in the United States, there was no significant difference in amplification efficiency or specificity. It is an effective exploration for the localization of reagents.
【學(xué)位授予單位】：四川大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2003
【分類(lèi)號(hào)】：D919

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前1條

1 張艷萍;DYS-714/723/650/713-plex通用引物嵌合復(fù)合擴(kuò)增及在法醫(yī)學(xué)實(shí)際檢案中的應(yīng)用性研究[D];山西醫(yī)科大學(xué);2010年

本文編號(hào)：2033749