| Centipedegrass (Eremochloa ophiuroides) belongs to the genus Eremochloa in Gramineae and is the only species used as turfgrass in the genus. Because of starting relatively late, the research of centipedegrass lagged far behind that of many other plants in constructing genetic linkage map, mapping QTL, genetic analysis of traits, estimating genetic diversity and constructing the core collection. In this study, the genetic diversity of centipedegrass germplasms was estimated and the core collection was also constructed. Based on this result, the first genetic linkage map was constructed by means of pseudo-test cross mapping strategy with Fi mapping population crossed by E102 and E092(1) that differed from morphological traits to genetic distance. The heredity of vegetative and reproductive traits was analyzed by the major gene and polygene mixed genetic model. On this basis, QTLs linked to these traits were also mapping in this map. The results were reported as follows:1 Analysis of genetic diversity and construction of core collection in centipedegrass1.1 Optimization of SRAP-PCR systemThe orthogonal design was used to optimize SRAP-PCR system with four factors (Mg2+, dNTP, primer and Taq polymerase) at three levels respectively. In addition, the concentration of template DNA was also selected. The results showed the optimized SRAP-PCR system for centipedegrass was:2μl 101xPCR buffer,60ng template DNA, Mg2+ 1.50mmol·L-1, dNTP260μmol·L-1, primer 0.25μmol·L-1,Taq DNA polymorphism 0.5U with total 20μl reaction solution. Each factors had different effect on the result of PCR. The concentration of Nig2+ had the greatest effect and Taq DNA polymorphism had the least effect on the result. The optimized SRAP-PCR system could be applied on molecular genetics research of centipedegrass.1.2 Analysis of genetic diversityThe genetic diversity of 148 centipedegrass germplasm was analyzed by using 27 SRAP primer combitions. The results showed:(1) Total 461 loci were amplified by 27 SRAP markers of which 397 were polymorphic with the percent of polymorphic loci as 86.12%. The Nei’s gene diversity was 0.2821 and Shannon diversity index was 0.4282 which indicated that a high level genetic diversity was detected in centipedegrass. (2) The genetic diversity of each population was calculated and the result showed that the order from high to low of genetic diversity level of these populations was Anhui, Hunan, Jiangsu, Guangdong, Henan, Zhejiang, Guizhou, Sichuan, Fujian, Chongqing, Jiangxi, Guangxi, USA and Hubei. (3) The result of genetic relationship among these populations showed that 25.59% and 74.71% genetic variance detected among and within populations, respectively. Gene flow was 1.4769 among populations which showed the genetic variance existed within populations. The result of UPGMA clustering showed that the Anhui and Jiangsu population were clustered together first and the population from Hubei was clustered lastly compared to other populations. Populations from neighboring province could priority cluster. (4) The result of UPGMA clustering from 148 materials showed these materials could not clustered strictly according to their geographic distance. But in each groups, there had a tendency that the materials clustered according to their geographic distance which showed there had certain correlation between the genetic distance and the geography distance of the centipedegrass germplasm.1.3 Construction of core collectionAccording to the result of clustering, clustering-grouping method, province grouping-grouping method and stepwise clustering method were used to screen the core collection of 148 centipedegrass germplasm. After compared the parameters which consisted of the number of polymorphic loci, percentage of polymorphic loci, observed the number of alleles, effective number of alleles, Nei’s gene diversity and Shannon’s information index of 12 sample groups,43 samples were chosen as core collection finally. T-test was performed between initial collection and core collection and the results showed that the core collection hold 29.7% samples of initial collection and the reserved rate of number of polymorphic loci and percentage of polymorphic loci reached 94.87%, the reserved rate of observed number of alleles, effective number of alleles, Nei’s gene diversity and Shannon’s information index were reached to 98.72%,100.91%,101.45%and 101.19% respectively. This result demonstrated that this core collection could well stand for initial collection.2 Genetic analysis for some important traits in centipedegrass2.1 Hybrids identification and selection of polymorphic SRAP primerThe hybrids status of centipedegrass was tested by using SRAP markers in this study. SRAP primer combinations which have polymorphic and paternal characteristic bands were selected by four parents (E022, E142, E102, E092(1)) from 400 primer combinations. Those SRAP markers with clear and stabilized paternal characteristic bands were used to test the hybrids status from two hybridization combinations. The results showed:162 (40.5%) polymorphic primer combinations were selection from 400 primer combinations and 288 polymorphic loci were amplified by these polymorphic primers. Polymorphic loci in each primer were 1.78. Fifty-one primer combinations with paternal characteristic bands were selected in which 6 primer combinations (303,297,283,180,332 and 358) were used to identify the hybrids status and 89 hybrids were true hybrids. These true hybrids could be used to construct the genetic linkage map and map QTL.2.2 Genetic analysis for vegetative traitsThe heredity of six vegetative traits, i.e. leaf length and width, internode length and diameter, height of turf and the color of stolon in F1 population were analyzed by the major gene and polygene mixed genetic model, in order to reveal the genetic mechanism of these traits. The results showed:(1) The widest variation is found in the leaf length that has a coefficient of variance (CV) of 23.39%, followed by the color of stolon, the variations of internode length, height of turf and internode diameter with each CV amounting to 21.96%, 21.89%,15.32% and 7.41% respectively. The coefficient variation of leaf width is the least, a mere 6.77%. (2) One major gene model with additive, partial dominance or over dominace (A-1) was the most suitable model for the leaf length and the heritability of major gene was 68.98%. One major gene model with negative entirely dominance (A-4) was the most suitable model for internode length and the heritability of major gene was 47.37%. Two major genes model with codominance (B-6) was the most suitable model for the height of turf and the heritability was 47.20%. The most suitable model of leaf width and internode diameter was no major gene model (A-0). The color of stolon was controlled by two addivive-dominance-epistasis major genes model (B-1) and the heritability of major genes is 99.23%.2.3 Genetic analysis for reproductive traits and correlation analysisThe heredity of reproductive characters, i.e. inflorescence density, reproductive branch height, inflorescence length and specule No. of each spike were analyzed by the major gene and polygene mixed genetic model, in order to reveal the genetic mechanism of these traits. The result showed:(1) The widest variation is found in the inflorescence density that has a coefficient of variance (CV) of 54.10%, followed by the variations of reproductive branch height, specule No. of each spike and inflorescence length with each CV amounting to 13.17%,21.89%,8.92% and 8.42% respectively. (2) One major gene model with negative entirely dominance (A-4) was the most suitable model for inflorescence density and the heritability of major gene was 60.71%. The reproductive branch height and inflorescence length were all controlled by two addivive-dominance-epistasis major genes model (B-1) and the heritability of major genes were 93.73% and 96.79% respectively. The most suitable model of specule No. of each spike was no major gene model (A-0). The result of correlation analysis showed:leaf length and width had very significant level which correlation coefficient 0.4178. Leaf length and height of turf also had very significant level which correlation coefficient was 0.6285. The reproductive branch height and inflorescence density, reproductive branch height and inflorescence length, reproductive branch height and height of turf had significant level which correlation coefficient were 0.3748,0.3562 and 0.4230, respectively.3 Construction of genetic linkage map in centipedegrass3.1 Construction genetic linkage map in centipedegrassBased on F1 hybrid population, the first genetic linkage map was constructed by means of pseudo-test cross mapping strategy using SRAP and EST-SSR (from wheat) markers. In female parent map,89 markers were located on 9 linkage groups with an average distance of 13.74cM between adjacent markers covering a total distance of 1209.08cM. The maximum distance between two markers was 38cM and the minimum distance was 1.1 cM. Sixteen gaps which distance exceeded 20cM were found in 3 linkage groups. In male parent map,81 markers were located on 12 linkage groups with an average distance of 15.48cM between adjacent markers covering a total distance of 1238.03cM. The maximum distance between two markers was 38cM and the minimum distance was 1.16cM. Eighteen gaps which distance exceeded 20cM were found in 8 linkage groups. These results could be used in construction high density genetic linkage map, mapping QTL, comparative mapping and marker assistance selection.3.2 Mapping QTL of some important traitsTwenty-seven putative QTLs were located in the genetic linkage map with WinQTLCart software by using composite interval mapping method. Except for color of stolon, the other 9 traits were all detected the QTLs. Eighteen QTLs were mapped in female parent map and 6 QTLs were mapped in male parent map. Six QTLs for leaf length were detected of which 3 were mapped in female parent map and 3 in male parent map. Five QTLs for leaf width were detected of which 4 in female parent map and 1 in male parent map. Four QTLs for internode length were detected and all mapped in female parent map. Only one QTL for internode diameter was mapped in female parent map. Two QTLs for height of turf were detected mapped in male and female parent map respectively. Four QTLs for inflorescence density were detected of which 2 mapped in female parent map and 2 in male parent map. Only one QTL for reproductive branch height was mapped in male parent map. Three QTLs for inflorescence length were detected of which 2 mapped in female parent map and 1 in male parent map. One QTL for specule No. of each spike was detected in male parent map. These results could provide theoretical basis for marker assistance selection and genetic improvement. |
PDF Full Text Request