Selection of high heterozygosity popcorn varieties in Brazil based on SSR markers

We analyzed genetic structure and diversity among eight populations of popcorn, using SSR loci as genetic markers. Our objectives were to select SSR loci that could be used to estimate genetic diversity within popcorn populations, and to analyze the genetic structure of promising populations with high levels of heterozygosity that could be used in breeding programs. Fifty-seven alleles (3.7 alleles per locus) were detected; the highest effective number of alleles (4.21) and the highest gene diversity (0.763) were found for the Umc2226 locus. A very high level of population differentiation was found (F(ST) = 0.3664), with F(ST) for each locus ranging from 0.1029 (Umc1664) to 0.6010 (Umc2350). This analysis allowed us to identify SSR loci with high levels of heterozygosity and heterozygous varieties, which could be selected for production of inbred lines and for developing new cultivars.     

Allelic reduction and genetic shift in the Canadian hard red spring wheat germplasm released from 1845 to 2004

Analysis of genetic diversity changes in existing gene pools of cultivated crops is important for understanding the impact of plant breeding on crop genetic diversity and developing effective indicators for genetic diversity of cultivated plants. The objective of this study was to assess genetic diversity changes in 75 Canadian hard red wheat (Triticum aestivum L.) cultivars released from 1845 to 2004 using 31 simple sequence repeats (SSRs) markers. A total of 267 SSR alleles were detected, and their allelic frequencies ranged from 0.01 to 0.97, with an average of 0.14. Significant allelic reduction was observed at only four SSR loci for the cultivars released from 1970 onwards. However, 51 alleles (about 19%) present in pre-1910 cultivars were undetected in cultivars released after 1990 and were spread over 27 SSR loci. The proportion of SSR variation accounted for by six breeding periods was 12.5%, by four ancestral families, 16.5%, and by eight breeding programs, 8.4%. The average genetic diversity measured by three different band-sharing methods did not change significantly among cultivars released from different breeding periods, breeding programs, and ancestral families. However, genetic shift was obvious in the cultivars released over the six breeding periods, reflecting well the various breeding efforts over years. These results clearly show the allelic reduction and genetic shift in the Canadian hard red spring wheat germplasm released over time. Consequently, more effort needs to be made to broaden the wheat breeding base and conserve wheat germplasm.     

Genetic structure of island populations of <Emphasis Type="Italic">Prunus lannesiana</Emphasis> var. <Emphasis Type="Italic">speciosa</Emphasis> revealed by chloroplast DNA,AFLP and nuclear SSR loci analyses

The wild flowering cherry Prunus lannesiana var. speciosa is highly geographically restricted, being confined to the Izu Islands and neighboring peninsulas in Japan. In an attempt to elucidate how populations of this species have established we investigated the genetic diversity and differentiation in seven populations (sampling 408 individuals in total), using three kinds of genetic markers: chloroplast DNA (cpDNA), amplified fragment length polymorphisms (AFLPs), and 11 nuclear SSR polymorphic loci. Eight haplotypes were identified based on the cpDNA sequence variations, 64 polymorphic fragments were scored for the AFLP markers, and a total of 154 alleles were detected at the 11 nuclear SSR loci. Analysis of molecular variance showed that among-population variation accounted for 16.55, 15.04 and 7.45% of the total detected variation at the cpDNA, AFLPs, and SSR loci, respectively. Thus, variation within populations accounted for most of the genetic variance for all types of markers, although the genetic differentiation among populations was also highly significant. For cpDNA variation, no clear structure was found among the populations, except that of the most distant island, although an “isolation by distance” pattern was found for each marker. Both neighbor-joining trees and structure analysis indicate that the genetic relationships between populations reflect geological variations between the peninsula and the islands and among the islands. Furthermore, hybridization with related species may have affected the genetic structure, and some genetic introgression is likely to have occurred.     

Microsatellite markers reveal high genetic diversity in date palm (<Emphasis Type="Italic">Phoenix dactylifera</Emphasis> L.) germplasm from Sudan

Genetic diversity in date palm germplasm from Sudan representing 37 female and 23 male accessions was investigated using 16 loci of microsatellite (SSR) primers. Eight female accessions from Morocco were included as reference material. The tested SSR markers showed a high level of polymorphism. A total of 343 alleles were detected at the 16 loci. The number of alleles per marker ranged from 14 to 44 with an average of 21.4 per locus. A high level of expected heterozygosity was observed among Sudan cultivars (0.841), Morocco cultivars (0.820) and male accessions (0.799). The results indicate that the genetic groups of the Sudan cultivars and/or males do not follow a clear geographic pattern. However, the morocco group showed significant differentiation in relation to the Sudan groups, as measured by F (ST) values and genetic distances. The effect of the methods of pollination and cultivar selection on the genetic structure was clearly detected by the weak clustering association that was observed for the majority of accessions originating from Sudan and Morocco as well. This suggests the need for further investigation on the genetic diversity of Sudanese date palm germplasm. A deeper insight will be revealed by a detailed analysis of populations originating from different geographic locations.     

Molecular characterization of oilseed rape accessions collected from multi continents for exploitation of potential heterotic group through SSR markers

Evaluation of the genetic diversity in conventional and modern rapeseed cultivars is essential for conservation, management and utilization of these genetic resources for high yielding hybrid production. The objective of this research was to evaluate a collection of 86 oilseed rape cultivars with 188 simple sequence repeat (SSR) markers to assess the genetic variability, heterotic group identity and relationships within and between the groups identified among the genotypes. A total of 631 alleles at 188 SSR markers were detected including 53 and 84 unique and private alleles respectively, which indicated great richness and uniqueness of genetic variation in these selected cultivars. The mean number of alleles per locus was 3.3 and the average polymorphic information content was 0.35 for all microsatellite loci. Unweighted Pair Group Method with Arithmetic Mean clustering and principal component analysis consistently divided all the cultivars into four distinct groups (I, II, III and IV) which largely coincided with their geographical distributions. The Chinese origin cultivars are predominantly assembled in Group II and showed wide genetic base because of its high allelic abundance at SSR loci while most of the exotic cultivars grouped into Group I and were highly distinct owing to the abundant private and unique alleles. The highest genetic distance was found between Group I and IV, which mainly comprised of exotic and newly synthesized yellow seeded (1728-1 and G1087) breeding lines, respectively. Our study provides important insights into further utilization of exotic Brassica napus accessions in Chinese rapeseed breeding and vice versa.     

Genetic variation and comparison of orchardgrass (Dactylis glomerata L.) cultivars and wild accessions as revealed by SSR markers

Orchardgrass is a highly variable, perennial forage grass that is cultivated throughout temperate and subtropical regions of the world. Despite its economic importance, the genetic relationship and distance among and within cultivars are largely unknown but would be of great interest for breeding programs. We investigated the molecular variation and structure of cultivar populations, compared the level of genetic diversity among cultivars (Baoxing, Anba, Bote, and Kaimo), subspecies (Dactylis glomerata ssp Woronowii) and advanced breeding line (YA02-116) to determine whether there is still sufficient genetic diversity within presently used cultivars for future breeding progress in China. Twenty individuals were analyzed from each of six accessions using SSR markers; 114 easily scored bands were generated from 15 SSR primer pairs, with an average of 7.6 alleles per locus. The polymorphic rate was 100% among the 120 individuals, reflecting a high degree of genetic diversity. Among the six accessions, the highest genetic diversity was observed in Kaimo (H = 0.2518; I = 0.3916; P = 87.3%) and 02-116 had a lower level of genetic diversity (H = 0.1806; I = 0.2788; P = 58.73%) compared with other cultivars tested. An of molecular variance revealed a much larger genetic variation within accessions (65%) than between them (35%). This observation suggests that these cultivars have potential for providing rich genetic resource for further breeding program. Furthermore, the study also indicated that Chinese orchardgrass breeding has involved strong selection for adaptation to forage production, which may result in restricted genetic base of orchardgrass cultivar.     

High-density Integrated Linkage Map Based on SSR Markers in Soybean

A well-saturated molecular linkage map is a prerequisite for modern plant breeding. Several genetic maps have been developed for soybean with various types of molecular markers. Simple sequence repeats (SSRs) are single-locus markers with high allelic variation and are widely applicable to different genotypes. We have now mapped 1810 SSR or sequence-tagged site markers in one or more of three recombinant inbred populations of soybean (the US cultivar ‘Jack’ × the Japanese cultivar ‘Fukuyutaka’, the Chinese cultivar ‘Peking’ × the Japanese cultivar ‘Akita’, and the Japanese cultivar ‘Misuzudaizu’ × the Chinese breeding line ‘Moshidou Gong 503’) and have aligned these markers with the 20 consensus linkage groups (LGs). The total length of the integrated linkage map was 2442.9 cM, and the average number of molecular markers was 90.5 (range of 70–114) for the 20 LGs. We examined allelic diversity for 1238 of the SSR markers among 23 soybean cultivars or lines and a wild accession. The number of alleles per locus ranged from 2 to 7, with an average of 2.8. Our high-density linkage map should facilitate ongoing and future genomic research such as analysis of quantitative trait loci and positional cloning in addition to marker-assisted selection in soybean breeding.Key words: EST-derived SSR marker, integrated linkage map, microsatellite marker, polymorphism information content     

Genetic identification of 91 poplar cultivars based on SSR markers

Accurate genetic identification and relationship analysis of poplar cultivars is necessary to establish commercial poplar plantations and select suitable breeding strategies. In this study, 91 poplar cultivars belonging to four sections (Aigeiros, Tacamahaca, Populus and Turanga) and inter/intra-sectional hybrids were genotyped using 18 polymorphic simple sequence repeat (SSR) markers. In total, 222 alleles were amplified with an average of 12.3 alleles per marker. The mean polymorphic information content and power of discrimination were 0.706 and 0.813, respectively. Five SSR markers (ORPM_103, ORPM_247, GCPM_1048, GCPM_1255 and LG_X_19) constituted a core fingerprint and were sufficient to identify all the tested cultivars. With some notable exceptions, cultivars of the same species generally clustered together in cluster (UPGMA) and ordination (PCO) analyses. Flow cytometry indicated that 11 poplar cultivars were triploid. Among these, seven had three alleles at some loci, suggesting that SSR markers could indicate the ploidy level to some extent. This study provides useful genetic information for the identification and protection of poplar cultivars in China and offers a guideline for the selection of poplar crossing parents based on ploidy level and genetic relationships.     

Identification of unique alleles and assessment of genetic diversity of rabi sorghum accessions using simple sequence repeat markers

Genetic diversity among 42 sorghum accessions representing landraces (19), advanced breeding lines (16), local cultivars (2) and release varieties (5) with 30 simple sequence repeat (SSR) markers revealed 7.6 mean number of alleles per locus showing 93.3% polymorphism and an average polymorphism information content of 0.78 which range from 0.22 (Xtxp12) and 0.91(Xtxp321). The average heterozygosity and effective number of alleles per locus were 0.8 and 6.65 respectively. Cluster analysis based on microsatellite allelic diversity clearly demarcated the accessions into ten clusters. A total of 24 unique alleles were obtained from seven SSR loci in 23 accessions in a size range of 110–380 bp; these unique alleles may serve as diagnostic tools for particular region of the genome of respective genotypes. Selected SSR markers from different linkage groups provided an accurate way of determining genetic diversity at the molecular level.     

Comparative analysis of microsatellite DNA polymorphism in landraces and cultivars of rice

Genetic polymorphisms of ten microsatellite DNA loci were examined among 238 accessions of landraces and cultivars that represent a significant portion of the distribution range for both indica and japonica groups of cultivated rice. In all, 93 alleles were identified with these ten markers. The number of alleles varied from a low of 3 or 4 at each of four loci, to an intermediate value of 9–14 at five loci, and to an extra-ordinarily high 25 at one locus. The numbers of alleles per locus are much larger than those detected using other types of markers. The number of alleles detected at a locus is significantly correlated with the number of simple sequence repeats in the targeted microsatellite DNA. Indica rice has about 14% more alleles than japonica rice, and such allele number differences are more pronounced in landraces than in cultivars. The indica-japonica differentiation component accounted for about 10% of the diversity in the total sample, and twice as much differentiation was detected in cultivars as in landraces. About two-thirds as many alleles were observed in cultivars as in landraces; another two-thirds of the alleles in the cultivar group were found in modern elite cultivars or parents of hybrid rice. The majority of the simple sequence repeat (SSR) alleles that were present in high or intermediate frequencies in landraces ultimately survived into modern elite cultivars and hybrids. The greater resolving power and the efficient production of massive amounts of SSR data may be particularly useful for germplasm assessment and evolutionary studies of crop plants.     

Genetic Diversity and Population Differentiation of Liaoning Weedy Rice Detected by RAPD and SSR Markers

Weedy rice refers to populations of usually annual Oryza species that diminish farmer income through reduction of grain yield and lowered commodity value at harvest. The genetic diversity and population genetic structure of weedy rice in Liaoning Province were studied by RAPD and SSR markers. The results indicate that the level of genetic diversity of Liaoning weedy rice is very low, with polymorphic loci being only 3.70% (RAPDs) and 47.62% (SSRs). On the other hand, high genetic differentiation was found among populations, in particular between two regions (Shenyang and Dandong), with Fst values of 0.746 (RAPDs) and 0.656 (SSRs), suggesting that more than two thirds of the genetic variation resides among regions. Combined with our investigations of cultural traditions, the low level of genetic diversity in Liaoning Province is attributed to its narrow genetic background enhanced by exchanges of cultivar seeds, whereas the high genetic differentiation between the two regions is most likely the result of different founding parents and gene flow from local rice varieties to weedy rice. The rice cultivars in the two regions are all local varieties and are different genetically. A comparison of the two marker systems demonstrates that SSR is more informative and powerful in terms of the assessment of genetic variability, although both RAPD and SSR provide useful genetic information on weedy rice.     

Analysis of genetic diversity and relationships among waxy maize inbred lines in Korea using SSR markers

Information regarding the genetic diversity and genetic relationships among elite inbred lines is necessary to improve new cultivars in maize breeding programs. In this study, genetic diversity and genetic relationships were investigated among 84 waxy maize inbred lines using 50 SSR markers. A total of 269 alleles were identified at all the loci with an average of 5.38 and a range between 2 and 13 alleles per locus. The gene diversity values varied from 0.383 to 0.923 with an average of 0.641. The cluster tree generated using the described SSR markers recognized two major groups at 32% genetic similarity. Group I included 33 inbred lines while group II included 51 inbred lines. The clustering patterns of most of the waxy maize inbred lines did not clearly agree with their source, pedigree or geographic location. The average GS among all inbred lines was 35.7 ± 10.8. Analysis of waxy maize inbred lines collected from Korea and China at 50 SSR loci revealed higher values of average number of alleles (4.9) and gene diversity (0.638) in Korean inbred lines as compared to Chinese inbred lines (3.5 and 0.563, respectively). The information obtained from the present studies would be very useful for maize breeding programs in Korea.     

Characterization of comparative genome‐derived simple sequence repeats for acanthopterygian fishes

Simple sequence repeats (SSRs) have become one of the most popular molecular markers for population genetic studies. The application of SSR markers has often been limited to source species because SSR loci are too labile to be maintained in even closely related species. However, a few extremely conserved SSR loci have been reported. Here, we tested for the presence of conserved SSR loci in acanthopterygian fishes, which include over 14 000 species, by comparing the genome sequences of four acanthopterygian fishes. We also examined the comparative genome‐derived SSRs (CG‐SSRs) for their transferability across acanthopterygian fishes and their applicability to population genetic analysis. Forty‐six SSR loci with conserved flanking regions were detected and examined for their transferability among seven nonacanthopterygian and 27 acanthopterygian fishes. The PCR amplification success rate in nonacanthopterygian fishes was low, ranging from 2.2% to 21.7%, except for Lophius litulon (Lophiiformes; 80.4%). Conversely, the rate in most acanthopterygian fishes exceeded 70.0%. Sequencing of these 46 loci revealed the presence of SSRs suitable for scoring while fragment analysis of 20 loci revealed polymorphisms in most of the acanthopterygian fishes. Population genetic analysis of Cottus pollux (Scorpaeniformes) and Sphaeramia orbicularis (Perciformes) using CG‐SSRs showed that these populations did not deviate from linkage equilibrium or Hardy–Weinberg equilibrium. Furthermore, almost no loci showed evidence of null alleles, suggesting that CG‐SSRs have strong resolving power for population genetic analysis. Our findings will facilitate the use of these markers in species in which markers remain to be identified.     

Comparative analysis of genetic diversity in the mangrove species Avicennia marina (Forsk.) Vierh. (Avicenniaceae) detected by AFLPs and SSRs

Avicennia marina is an important mangrove species with a wide geographical and climatic distribution which suggests that large amounts of genetic diversity are available for conservation and breeding programs. In this study we compare the informativeness of AFLPs and SSRs for assessing genetic diversity within and among individuals, populations and subspecies of A. marina in Australia. Our comparison utilized three SSR loci and three AFLP primer sets that were known to be polymorphic, and could be run in a single analysis on a capillary electrophoresis system, using different- colored fluorescent dyes. A total of 120 individuals representing six populations and three subspecies were sampled. At the locus level, SSRs were considerably more variable than AFLPs, with a total of 52 alleles and an average heterozygosity of 0.78. Average heterozygosity for AFLPs was 0.193, but all of the 918 bands scored were polymorphic. Thus, AFLPs were considerably more efficient at revealing polymorphic loci than SSRs despite lower average heterozygosities. SSRs detected more genetic differentiation between populations (19 vs 9%) and subspecies (35 vs 11%) than AFLPs. Principal co-ordinate analysis revealed congruent patterns of genetic relationships at the individual, population and subspecific levels for both data sets. Mantel testing confirmed congruence between AFLP and SSR genetic distances among, but not within, population comparisons, indicating that the markers were segregating independently but that evolutionary groups (populations and subspecies) were similar. Three genetic criteria of importance for defining priorities for ex situ collections or in situ conservation programs (number of alleles, number of locally common alleles and number of private alleles) were correlated between the AFLP and SSR data sets. The congruence between AFLP and SSR data sets suggest that either method, or a combination, is applicable to expanded genetic studies of mangroves. The codominant nature of SSRs makes them ideal for further population-based investigations, such as mating-system analyses, for which the dominant AFLP markers are less well suited. AFLPs may be particularly useful for monitoring propagation programs and identifying duplicates within collections, since a single PCR assay can reveal many loci at once. Received: 3 October 2000 / Accepted: 19 February 2001     

Analysis of genetic diversity and population structure of peanut cultivars and breeding lines from China,India and the US using simple sequence repeat markers

Cultivated peanut is grown worldwide as richsource of oil and protein. A broad genetic base is needed for cultivar improvement. The objectives of this study were to develop highly informative simple sequence repeat(SSR)markers and to assess the genetic diversity and population structure of peanut cultivars and breeding lines from different breeding programs in China, India and the US. A total of 111 SSR markers were selected for this study, resulting in a total of 472 alleles. The mean values of gene diversity and polymorphic information content(PIC) were 0.480 and 0.429, respectively.Country-wise analysis revealed that alleles per locus in three countries were similar. The mean gene diversity in the US,China and India was 0.363, 0.489 and 0.47 with an average PIC of 0.323, 0.43 and 0.412, respectively. Genetic analysis using the STRUCTURE divided these peanut lines into two populations(P_1, P_2), which was consistent with the dendrogram based on genetic distance(G_1, G_2) and the clustering of principal component analysis. The groupings were related to peanut market types and the geographic origin with a few admixtures. The results could be used by breeding programs to assess the genetic diversity of breeding materials to broaden the genetic base and for molecular genetics studies.     

Assessment of Genetic Diversity in <Emphasis Type="Italic">Secale cereale</Emphasis> Based on SSR Markers

The primary aim of this study was to estimate genetic diversity among Secale cereale L. accessions using 22 previously published simple sequence repeat (SSR) markers. The plant material included 367 rye accessions comprising historical and contemporary cultivars, cultivated materials, landraces, and breeding strains from the Polish breeding company Danko. The studied accessions represented a wide geographical diversity. Several methods were employed to analyze genetic diversity among the Secale cereale L. accessions and to determine population structure: principal coordinate analysis (PCoA), neighbor-joining (NJ), and Bayesian clustering. We also defined a core collection of 25 rye accessions representing over 93 % of SSR alleles. The results of these analyses showed that accessions from the rye gene bank are clearly divergent in comparison with materials received directly from European breeding companies. Our findings suggest also that the genetic pool of current rye cultivars is becoming narrower during breeding processes. The selected panel of SSR markers performed well in detection of genetic diversity patterns and can be recommended for future germplasm characterization studies in rye.     

Automated sizing of fluorescent-labeled simple sequence repeat (SSR) markers to assay genetic variation in soybean

 Simple Sequence Repeat (SSR) allele sizing provides a useful tool for genotype identification, pedigree analysis, and for estimating genetic distance between organisms. Soybean [Glycine max (L.) Merr.] cultivars are identified for Plant Variety Protection (PVP) purposes by standard pigmentation and morphological traits. However, many commercial soybeans arise from a limited number of elite lines and are often indistinguishable based on these traits. A system based on SSR markers would provide unique DNA profiles of cultivars. Fluorescent labeling of alleles combined with automated sizing with internal size standards in each gel lane was used as an alternative to standard [³²P] labeling to assess genetic variability in soybean. Allelic frequencies at 20 SSR loci were determined in 35 soybean genotypes that account for greater than 95% of the alleles in North American soybean cultivars based upon pedigree analysis. An average of 10.1 alleles per locus (range: 5–17), with a mean gene diversity of 0.80 (range: 0.50 to 0.87) were observed at the 20 SSR loci. The 20 loci successfully distinguished modern soybean cultivars that are identical for morphological and pigmentation traits, as well as 7 soybean genotypes reported to be indistinguishable using 17 RFLP probes. Pedigrees of 7 cultivars were studied to estimate stability of SSRs in soybean across generations. Of the 7 pedigrees 6 had one locus in the progeny with an allele(s) that was not present in either parent. These new alleles are most likely the result of mutation. The mutation rate of SSR alleles in soybean was similar to that reported in humans. To avoid difficulty associated with mutation, DNA fingerprint data should be determined from the bulk of 30-50 plants of a cultivar. Received: 24 March 1997 / Accepted: 4 April 1997     

Evaluation of Genetic Diversity in Chinese Wild Apple Species Along with Apple Cultivars Using SSR Markers

China, one of the primary centers of genetic diversity for the genus Malus, is very rich in wild apple germplasm. In this study, genetic diversity in 29 Malus accessions, including 12 accessions from 7 Chinese Malus species, 4 Chinese landraces, and 13 introduced apple cultivars, was assessed using a set of 19 single-locus simple sequence repeat (SSR) markers distributed across all 17 linkage groups of the apple genome. The number of alleles detected at each locus ranged from 2 to 11, with an average of 5.3 per SSR marker. In some accessions, 16 unique alleles were identified. Ten out of these 16 unique alleles (62.5%) were detected exclusively in wild species, indicating that these Chinese wild apple species have considerable genetic diversity and can be used in breeding programs to increase the genetic diversity of apple cultivars. Using 19 SSRs, an unweighted pair-group method with arithmetic average cluster analysis was conducted, and the resulting dendrogram revealed that all cultivars, except for E??peMeBckoe, were clustered together in the same group. The Russian cultivar E??peMeBckoe was closely related to the Chinese crabapple Baihaitang (M. prunifolia), with a high similarity coefficient value of 0.94. Of the two M. sieversii accessions used, one accession showed a close relationship to apple cultivars, while the other accession was closely related to wild apple species, suggesting the presence of a wider genetic diversity in Chinese M. sieversii species. The influence of SSR marker selection on genetic diversity analysis in this Malus collection was also discussed.