利用重组自交系群体构建番茄AFLP遗传连锁图谱

Construction AFLP Genetic Linkage Map of Tomato Using Recombinant Inbred Lines(RILs)Population

以普通栽培番茄(Solanum lycopersicum)99165-30为母本,野生多毛番茄(Solanum habrochaites)LA1777为父本进行杂交,通过单粒传得到了含有80个F5︰6家系的重组自交系分离群体,利用荧光AFLP分子标记技术构建番茄分子遗传连锁图谱。AFLP标记采用MseⅠ和EcoRⅠ两种内切酶及荧光标记(IRD-700或IRD-800)的E+3和非荧光标记的M+3引物组合进行选择性扩增,扩增结果经95℃预变性后在6%变性聚丙烯酰胺凝胶上电泳2.5h,运用LICOR公司的NEN Global Edition IR2 DNA Analyzer(Model 5200 LI-COR Biosciences,Lincoln,NE)荧光扫描检测DNA多态性。对RILs群体中产生分离的274个AFLP标记运用JoinMap3.0软件分析,得到一张番茄分子遗传连锁图谱,图谱总长度为662cM,共包括18个主要连锁群,125个多态性分子标记。每条连锁群上的标记数在3～22个之间,连锁群的长度在14.0～58.0cM的范围内,平均图距在2.27～13.3cM。总平均距离5.3cM,本研究中构建的番茄永久遗传图谱,为番茄分子辅助育种及重要农艺性状的定位奠定了基础。

A genetic linkage map of tomato was constructed using a RILs （recombinant inbred lines） population of 80 individuals which was developed by crossing Solanum lycopersicum 99165-30 and Solanum habrochaites LA1777 through single-seed descent （SSD） . AFLPs were generated by the use of restriction enzymes EcoR I in combination with either Mse I. Pre-amplification was carded out using primers corresponding to EcoR I and Mse I adaptors with no selective base. Selective amplifications were performed using IRD700 or IRD800 labeled EcoR I primers and non-labeled Mse I primers. The resulting products were denatured in formamide at 95℃ and separated by electrophoresis 2.5 h on 6% polyacrylamide gel using IR2 DNA Analyzer （Model 5200 LI-COR Biosciences, Lincoln, NE） . The segregation of each marker and linkage analysis was done using the program JoinMap version 3.0. With 22 primer pairs, a total of 247 parental polymorphic bands were detected and 125 used for mapping, the total map length was 662 cM, consisted of 18 linkage groups, number of markers in the linkage groups varied from 3 to 22, the length of linkage groups were from 14.0 cM to 58.0 cM and mean marker interval distance from 2.27 cM to 13.3 cM individually, and a mean marker interval distance of 5.3 cM between markers. The map developed in the present study could be used for genetic mapping and molecular marker assisted breeding and quantitative trait locus mapping of tomato.