Large-scale Development And Application Of EST-SSR Markers Of Liriodendron

Magnoliaceae is a primitive group of angiosperms, and plays an important role in plantsystem evolution. Little progress has been achieved on the molecular genetics research ofMagnoliaceae plants due to the lack of suitable DNA markers. In recent years, although a numberof SSR markers were developed from EST database of Liriodendron and their cross-generatransferability was also reported, however, the number of SSR markers in Magnoliaceae is stilllimited. Thus, large-scale development EST-SSR primers of Liriodendron and detection ofcross-genera transferability is of practical importance. In this paper, we performed thetranscriptome sequencing of L. chinensis, and developed a number of SSR primers from the ESTdatabase of L. chinensis and L. tulipifera. The author also evaluated their application prospects ofthis set of SSR markers in the detection of population genetic diversity in L. chinensis as well astheir cross-genera transferability in Magnoliaceae. The main results are as follows:Large-scale development of EST-SSR primers of Liriodendron The transcriptomesequencing of petals and leaves of L. chinensis was performed using the second-generationsequencing technologies. The total sequences were up to17G bp, and129,097ESTs were gotthrough data filtering and splicing. Ultimately2,001pairs of SSR primers were developed fromEST database of L.chinensis, and861pairs of EST-SSR primers from EST database ofL.tulipifera.The detection of genetic diversity in a natural population of L. chinensis usingEST-SSR markers of L.tulipifera. The polymorphism of861pairs of primers was evaluated bytaking6genotypes of L.tulipifera as well as6genotypes of L.chinensis.as templates for PCRamplification. Of them,671pairs of primers amplified in Liriodendron, and372pairs of primerswere amplified polymorphism, with a polymorphic primers ratio of55.44%.26pairs of primerswere applied to detect the genetic diversity in a natural population of L.chinensis fromMaoershan Reserve, Guilin, Guangxi.18pairs of primers amplified polymorphic products. Thetotal allele number was104, with average allele number of5.78per locus. The effective allelenumbers ranged from2.96to5.86, and the average effective allele numbers was4.16. Theaverage Shannon diversity index and the average expected heterozygosity were1.5447,0.7632respectively, indicating that this natural population maintains high genetic diversity. It alsoshowed that these set of SSR primers could have extensive application prospect.The analysis of polymorphism and cross-genera transferability of EST-SSR primers ofL.chinensis The polymorphism of192pairs of primers were detected by taking6genotypes of L.tulipifera as well as6genotypes of L.chinensis.as templates for PCR amplification.120pairsof primers amplified successfully in L.chinensis, and44pairs of primers were amplifiedpolymorphism, with a polymorphic primers ratio of.36.7%.107pairs of primers amplifiedsuccessfully in L.tulipifera.34pairs of primers were amplified polymorphism, with apolymorphic primers ratio of31.8%. The within-genus transferability rate of EST-SSR primerswas89.2%.36pairs of highly polymorphic primers were applied to detect their cross-generatransferability. The transferability rates in Manglietia, Magnolia, Michelia, Kmeria, TalaumaJuss., Parakmeria Hu, Tsoongiodendron Chun, Manglietiastrum Law, Paramichelia Hu were22.2%to50.0%,27.8%to44.4%,36.1%to52.8%,33.3%,44.4%,55.6%,52.8%,50.0%,41.7%respectively. These results indicated that these set of EST-SSR primers could be applied in otherMagnoliaceae plants.