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  本文關(guān)鍵詞： 單性花山葡萄 遺傳圖譜 SSR SRAP　出處：《沈陽農(nóng)業(yè)大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：山葡萄(Vitis amurensisRupr.)具有很強(qiáng)的抗寒性與抗病性,是培育抗寒、抗病新品種及砧木育種的寶貴植物材料。本研究選用雌能花山葡萄'左山一'和雄性花山葡萄'043'為親本雜交創(chuàng)建了 F1代作圖群體,采用SSR和SRAP兩種分子標(biāo)記技術(shù),構(gòu)建了山葡萄分子遺傳圖譜,為后期實現(xiàn)與性別相關(guān)基因位點的定位提供理論依據(jù),也為抗寒抗病等育種工作打下良好基礎(chǔ),主要研究結(jié)果如下:1.應(yīng)用SSR-PCR技術(shù)篩選出兩對具有父母本特異性條帶的SSR引物,并對148株雜交后代進(jìn)行雜種真實性鑒定,共得到142株真雜種,真雜種率為95.9%。并創(chuàng)建了由142個真雜種組成的F1代作圖群體。2.建立并優(yōu)化了 SSR-PCR和SRAP-PCR反應(yīng)體系。兩者最佳反應(yīng)體積都是16 μ 1,前者反應(yīng)體系中,模板DNA含量為1.5ng,Mg2+濃度為1.0mmol/L,上下游引物濃度均為0.5mmol/L,dNTPs濃度為0.6mmol/L,TaqDNA聚合酶含量為0.5U。后者反應(yīng)體系中,模板DNA濃度為0.9ng,Mg2+濃度為0.8mmol/L,上下游Primer濃度均為0.6 μ mol/L,dNTPs濃度為1.2mmol/L,Taq DNA聚合酶含量為0.6U,其余部分是滅菌后雙蒸水。并驗證了建立的SSR-PCR和SRAP-PCR反應(yīng)體系的可靠性,表明倆個最佳體系可用于雜種鑒定及山葡萄遺傳圖譜構(gòu)建等后續(xù)試驗。3.得到153對至少在一個親本上表現(xiàn)為雜合,在雙親間表現(xiàn)出多態(tài)性且在后代產(chǎn)生分離的SSR引物。其中在父本上得到96個分離位點,在母本上得到107個分離位點。用167對多態(tài)性良好的SRAP引物以作圖群體擴(kuò)增,一共得到258個分離位點。4.構(gòu)建得到雌能花山葡萄'左山一'的分子遺傳圖譜,包括19個連鎖群,覆蓋總圖距為1551.6cM,連鎖群平均長度為81.7cM,位點之間平均距離為5.6cM,每個連鎖群平均包含14.6個,位點位點間最短距離是0.1cM;雄性花山葡萄'043'的分子遺傳圖譜,包括19個連鎖群,覆蓋總圖距為1540.9cM,連鎖群平均長度為81.1cM,位點之間平均距離為5.3cM,每個連鎖群平均包含15.3個位點,位點間最短距離是0.3cM。
[Abstract]:Vitis amurensis Rupr.) has strong cold resistance and disease resistance. In this study, the female grape 'Zuoshan' and the male 'Zaoshan' were used as parents to create the F1 generation mapping population for new resistant varieties and valuable plant materials for rootstock breeding. The molecular genetic map of Vitis vinifera was constructed by using SSR and SRAP molecular markers, which provided the theoretical basis for the localization of sex-related gene loci in the later stage. It also laid a good foundation for cold resistance and disease resistance breeding. The main results are as follows: 1. Two pairs of SSR primers with parental specific bands were screened by using SSR-PCR technique. A total of 142 true hybrids were obtained by identifying the hybrids of 148 hybrids. The true hybrid rate is 95.9.The F1 generation mapping population, which consists of 142 true hybrids, has been established and optimized. The optimum reaction volume of SSR-PCR and SRAP-PCR was 16 渭 1. In the former reaction system, the concentration of template DNA was 1.0 mmol / L and the concentration of upstream and downstream primer was 0.5 mmol / L. The concentration of dNTPs was 0.6 mmol / L, the concentration of Taq DNA polymerase was 0.5 U.In the latter reaction system, the concentration of template DNA was 0.9ng. The concentration of Mg2 was 0.8 mmol / L, and the concentration of upstream and downstream Primer was 0.6 渭 mol / L, the concentration of dNTPs was 1.2 mmol / L. The content of Taq DNA polymerase was 0.6U, and the rest was double-steamed water after sterilization. The reliability of the established reaction system of SSR-PCR and SRAP-PCR was verified. The results showed that the two optimal systems could be used in further experiments such as hybrid identification and genetic map construction of Vitis vinifera. 153 pairs were found to be heterozygotes on at least one parent. The SSR primer showed polymorphism between parents and produced segregation in offspring. 96 segregation loci were obtained on male parent. 107 segregation loci were obtained on the female parent, and 167 pairs of polymorphic SRAP primers were used to map the population amplification. A total of 258 segregation loci were obtained. The molecular genetic map of 'Zuoshan' was constructed, including 19 linkage groups, covering a total map distance of 1551.6 cm. The average length of linkage group was 81.7 cm, the average distance between loci was 5.6 cm, the average length of linkage group was 14.6, and the shortest distance between loci was 0.1 cm. The molecular genetic map of male Grape Grape, including 19 linkage groups, covers a total map of 1540.9 cm, and the average length of linkage group is 81.1 cm. The average distance between loci was 5.3 cm, and the average number of loci in each linkage group was 15.3, and the shortest distance between loci was 0.3 cm.
【學(xué)位授予單位】：沈陽農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：S663.1

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本文編號：1474154