Dominant Amplified Fragment Length Polymorphism Research Articles

Landscape complexity has limited effects on the genetic structure of two arable plant species, Adonis aestivalis and Consolida regalis

SummaryThe agricultural landscape of central Europe has changed dramatically in recent decades due to intensified cultivation, bringing many of its characteristic species to the brink of extinction. We investigated whether landscape structure affects the genetic structure and diversity of remnant populations of the two arable plant species Adonis aestivalis and Consolida regalis. We used dominant amplified fragment length polymorphism markers (AFLPs) and compared populations from six regions of 5 km² in central Germany. These regions represent two different classes of landscape structural complexity: intensively used, homogeneous landscapes (>95% of area covered by arable land with low extent of field margins) or heterogeneous regions (<60% of area covered by arable land with large extent of field margins). Contrary to expectations, within‐population diversity levels did not significantly differ between homogeneous and heterogeneous landscapes. No significant isolation‐by‐distance was found for either species, regardless of landscape structure, and genetic structures may still mirror more continuous conditions before large‐scale restructuring commenced in Germany's arable landscapes from the 1950s onwards. These results suggest that current landscape complexity, as such, is not as important for local‐level genetic structure for the species studied. Thus, homogenised and intensively used landscapes may also be important for the conservation of arable plant diversity and should not be neglected.

Genome mapping in F1 population of crossbred Italia and Mercan grape varieties: establishment of AFLP and SSR linkage groups towards significant morphological characters and fungal diseases.

This research aims at the establishment of a genome map for F 1 population developed by crossbreeding of Italia and Mercan grape varieties. A co-dominant marker SSR (Simple Sequence Repeats) and a dominant marker AFLP (Amplified Fragment Length Polymorphism) were used in the study. Double-pseudotestcross mapping technique was used for genome mapping. The study analyzed 60 plants selected from the F 1 population as well as the parental grape varieties, which differed in regards to resistance to diseases and some morphological characteristics. Amplification products were evaluated as “available” and “not available” at the end of silver staining for SSR and radioactive marking for AFLP. Polymorphic loci were determined according to their segregation ratios ranging between 1: 1 and 3: 1 at the end of X 2 tests. 3.0 LOD value was used in Mapmaker/Exp 3.0 package program for mapping. Two separate genetic linkage maps (maternal and paternal) were observed which included 6 and 1 linkage groups, respectively. Linkage of the loci located on the linkage groups to the observed diseases and morphological characters were analyzed with regression and variance analyses. A total of 818 primer pairs were tested on the two parents (Italia and Mercan), 60 F 1 (Italia × Mercan population) and two reference grape cultivar (Cabarnet Sauvignon and Merlot), successfully amplifying 112 markers. When the resistance traits to fungal diseases were analyzed during the study, no markers related with resistance to Botrytis cinerea and downy mildew could be found. However, it was found that the markers (VrZAG29a, VrZAG29c) related with the control of resistance to powdery mildew were on a locus which was in linkage group. Keywords: Vitis vinifera L., grape, SSR, AFLP, mapping African Journal of Biotechnology Vol. 9(36), pp. 5879-5886, 6 September, 2010

Landscape, population structure and genetic diversity ofStomoxys calcitrans

To investigate whether different landscapes could affect genetic diversity and structure of the cosmopolitan diptera Stomoxys calcitrans, populations from Gabon and southern France were studied using dominant amplified fragment length polymorphism (AFLP) markers. Gabon is characterized by a forested closed landscape, and southern France by an open Mediterranean landscape. The genetic diversity between Gabon and France populations did not differ significantly (P > 0.05). Contrary to our expectation, this study shows a moderate level of genetic differentiation between these two distant countries (Fst = 0.0979) and a low genetic structure among Gabonese and French populations (Fst = 0.0291 and 0.0275 respectively). This result could indicate the capacities of S. calcitrans populations to sustain a high level of gene flow, despite geographic distance and isolation.

Genetic diversity of relictual and endangered plant Abies ziyuanensis (Pinaceae) revealed by AFLP and SSR markers

Abies ziyuanensis is a highly endangered fir species endemic to South China. Unlike other Abies species that are distributed in areas with cold climates, A. ziyuanensis is restricted to several isolated island-like localities at subtropical mountains. In this study, we used dominant amplified fragment length polymorphism (AFLP) and co-dominant simple sequence repeats (SSR) markers to infer the genetic structure of A. ziyuanensis. Seven populations consisting of 139 individuals were sampled across their whole distribution. A. ziyuanenesis has a relatively low level of genetic variation, with a mean genetic diversity per population (He) of 0.136 (AFLP) and 0.337 (SSR), which is lower than that of other reported endemic species based on the same kind of marker. We observed high population differentiation, with Gst = 0.482 (AFLP) and Fst = 0.250 (SSR), among the seven populations. AMOVA also detected significant differentiation among populations (Phist (AFLP) = 0.550 and Phist (SSR) = 0.289) and among regions (Phict (AFLP) = 0.139 and Phict (SSR) = 0.135) in both marker types. Both ongoing evolutionary forces (e.g., genetic drift resulting from small population size) and historical events (e.g., population contraction and fragmentation during and after the Quaternary glacial cycles) may have contributed to the genetic structure in A. ziyuanensis.

QTL Analysis for Resistance to Phytophthora capsici in Pepper Using a High Density SSR-based Map

A segregating doubled haploid (DH) population (n = 96) was developed by anther culture of an F1 plant crossed between susceptible (‘Manganji’) and resistant (‘Criollo de Morelos 334’) lines of pepper (Capsicum annuum L.) to conduct a genetic analysis of resistance to Phytophthora rot caused by Phytophthora capsici. In order to perform a quantitative trait locus (QTL) analysis, we constructed a high density simple sequence repeat (SSR)-based map with a total length of 878 cM. Sixteen linkage groups (LGs) and 118 SSR markers were located using the 626 SSR markers that we previously developed. Resistance was evaluated in two root inoculation tests. Interval mapping for the resistance to P. capsici detected a common major QTL in the duplicate tests and a minor QTL specific to the first test. The major QTL was located on LG15 and flanked with an SSR marker, CAMS420. In addition, seven SSR markers were located within 21 cM intervals from the peak of this QTL. In contrast, the QTL on LG3 was detected with small effects in the first test, the nearest marker was a dominant amplified fragment length polymorphism (AFLP) marker, and the QTL was surrounded by eight SSR markers within a distance of 10 cM. Since some of the linkage markers for agriculturally valuable traits cannot detect polymorphism within breeding populations in C. annuum, the present linkage markers may widen the choice in marker-assisted selection in breeding programs for Phytophthora rot resistant pepper cultivars.

Genetic structure of the forest pest Hylobius abietis on conifer plantations at different spatial scales in Europe

The distribution of genetic variation within and among 20 European sites infested by the large pine weevil, Hylobius abietis, was analysed using dominant amplified fragment length polymorphism markers. Analysis of molecular variance was performed at the European, regional and local scales. Most of the genetic variability was found within rather than among populations and the global fixation index averaged over loci was low (0.07). We found no evidence of genetic drift, even in relatively isolated sites. This genetic pattern tends to confirm the high dispersal ability of the weevil and the influence of human-mediated expansion of its range through conifer plantations across Europe since the 19th century. Assignment tests demonstrated that the regional forest is a pertinent geographic scale for defining populations in the large pine weevil. Testing the potential influence of the larval host-plant identity (Scot Pine vs Norway Spruce) on the genetic structure revealed a weak but significant effect in two of the three regions tested (in Ardèche and in Limousin but not in Finland). One locus varied with host-plant use in the two French regions, indicating a potential role in host-plant adaptation. However, host-race formation is not observed in H. abietis; we discuss this result in the light of our current knowledge of this insect's biology. Altogether, this study shows that the use of different host plants for development does not constitute a strong barrier to gene flow for H. abietis and confirms the high dispersal ability of this forest pest.

Genetic diversity and differentiation in Eryngium alpinum L. (Apiaceae): comparison of AFLP and microsatellite markers.

Genetic diversity and structure of 12 populations of Eryngium alpinum L. were investigated using 63 dominant amplified fragment length polymorphism (AFLP) and seven codominant microsatellite (48 alleles) markers. Within-population diversity estimates obtained with both markers were not correlated, but the microsatellite-based fixation index Fis was correlated with both AFLP diversity indices (number of polymorphic bands and Nei's expected heterozygosity). Only AFLP diversity indices increased with the size of populations, although they did not significantly differ among them (Kruskall-Wallis test). The discrepancy between AFLPs and microsatellites may be explained by a better coverage of the genome with numerous AFLPs, the higher mutation rates of microsatellites or the absence of significant difference among within-population diversity estimates. Genetic differentiation was higher with AFLPs (theta=0.40) than with microsatellites (theta=0.23), probably due to the higher polymorphism of microsatellites. Thus, we considered global qualitative patterns rather than absolute estimates to compare the performance of both types of markers. On a large geographic scale, the Mantel test and multivariate analysis showed that genetic patterns were more congruent with the spatial arrangement of populations when inferred from microsatellites than from AFLPs, suggesting higher homoplasy of AFLP markers. On a small spatial scale, AFLPs managed to discriminate individuals from neighboring populations whereas microsatellites did not (multivariate analysis), and the percentage of individuals correctly assigned to their population of origin was higher with AFLPs than with microsatellites. However, dominant AFLPs cannot be used to study heterozygosity-related topics. Thus, distinct molecular markers should be used depending on the biological question and the geographical scale investigated.

Merging Carrot Linkage Groups based on Conserved Dominant AFLP Markers in F2 Populations

Markers were placed on linkage groups, ordered, and merged for two unrelated F2 populations of carrot (Daucus carota L.). Included were 277 and 242 dominant Amplified fragment-length polymorphism (AFLP) markers and 10 and eight codominant markers assigned to the nine linkage groups of Brasilia × HCM and B493 × QAL F2 populations, respectively. The merged linkage groups were based on two codominant markers and 28 conserved dominant AFLP markers (based upon sequence and size) shared by both populations. The average marker spacing was 4.8 to 5.5 cM in the four parental coupling phase maps. The average marker spacing in the six merged linkage groups was 3.75 cM with maximum gaps among linkage groups ranging from 8.0 to 19.8 cM. Gaps of a similar size were observed with the linkage coupling phase maps of the parents, indicating that linkage group integration did not double the bias which comes with repulsion phase mapping. Three out of nine linkage groups of carrot were not merged due to the absence of common markers. The six merged linkage groups incorporated similar numbers of AFLP fragments from the four parents, further indicating no significant increase in bias expected with repulsion phase linkage. While other studies have merged linkage maps with shared AFLPs of similar size, this is the first report to use shared AFLPs with highly conserved sequence to merge linkage maps in carrot. The genome coverage in this study is suitable to apply quantitative trait locus analysis and to construct a cross-validated consensus map of carrot, which is an important step toward an integrated map of carrot.

Genetic structuring of the stream lily Helmholtzia glaberrima (Philydraceae) within Toolona Creek, south-eastern Queensland

The distribution of genetic variation among five isolated sites of the riparian species Helmholtzia glaberrima (J.D.Hook) was examined in Toloona Creek (28�13′S, 153�07′E) by using dominant amplified fragment length polymorphism (AFLP) markers. From the 137 fragments assessed, analysis of molecular variance (AMOVA) showed that most genetic variability occurred within sites (68%), although high (32%) variation also occurred among sites. Highly significant pairwise θ estimates among all sampled sites suggest that gene flow is restricted in H. glaberrima. Levels of within-site diversity were intermediate and significantly different across the sampled sites. Significant levels of linkage disequilibrium were detected at all sites except TC3. Differences in linkage disequilibrium and genetic diversity among the sites suggest that sites may have been founded by different numbers of colonists. Mantel tests found no correlation between geographic and genetic distance and significant levels of linkage disequilibrium were detected at the total site level, supporting a non-equilibrium model of population structure. The observed pattern of non-equilibrium population structure and genetic variation in H. glaberrima are best explained by a classical metapopulation model.

Genetic mapping of the evergrowing gene in peach [Prunus persica (L.) Batsch

In temperate locations, terminal apices on evergrowing (also called evergreen) peach trees keep growing in winter until killed by low temperatures, while the lateral buds go into dormancy. A recessive allele of a single gene (evergrowing or evg) controls this trait in peach. The amplified fragment length polymorphism (AFLP) technique and bulked segregant analysis were applied to construct a local genetic linkage map for the evg gene from the cross Empress op op dwarf x Evergrowing (P.I. 442380). This map, comprising nine AFLP markers and the evg locus, covers a total genetic distance of 79.3 cM. Four dominant AFLP markers (EAT/MCAC, ETT/MCCA2, EAT/MCTA, and ETT/MACC) were linked to the evg locus at distances of 1, 5.3, 6.7, and 11.7 cM, respectively. EAT/MCAC and EAT/MCTA were converted into polymorphic sequence-tagged sites. Microsatellite markers in the evg region were developed from peach bacterial artificial chromosome (BAC) clones that hybridized to the AFLP marker fragments. Using three microsatellite anchor markers (pchgms12, pchgms17, and pchgms19), the local genetic linkage map was integrated into one minor linkage group of a previously constructed peach rootstock genetic linkage map. Three AFLP markers from the rootstock genetic linkage map were found linked to the evg locus.

Amplified fragment length polymorphism (AFLP) markers reveal extra-pair parentage in a bird species: the bluethroat (Luscinia svecica).

We tested the use of amplified fragment length polymorphism (AFLP) to assess the frequency of extra-pair parentage in a bluethroat (Luscinia svecica namnetum) population. Thirty-six families totalling 162 nestlings were analysed. Using a combination of three primer pairs, we reached an exclusion probability of 93% for the population. This probability can reach 99% considering families independently. We revealed that extra-pair fertilizations are very common: 63.8% of all broods contain at least one extra-pair young, totalling 41.9% of all young analysed. However, with the technique and the three primer pairs used it was not possible to attribute the parentage exclusions to extra-pair paternity, maternity or both. As brood parasitism has never been reported in this species, it seems likely that the exclusions are due to extra-pair males. This study shows that dominant AFLP markers can be useful for studying the mating system of taxa for which no microsatellite primers are available. This technique allows the approximate estimation of parentage exclusions despite the fact that it is not possible to know which parent has to be excluded.

Construction of a genetic linkage map and identification of AFLP markers for resistance to root-knot nematodes in peach rootstocks

A genetic linkage map for peach (Prunus persica (L.) Batsch) rootstocks has been constructed using amplified fragment length polymorphism (AFLP) markers in 55 F2individuals derived from the cross Lovell x Nemared. From 21 different primer combinations, a total of 169 AFLP markers were scored, of which, 153 were assigned to 15 linkage groups covering 1297 centimorgans (cM) of the peach nuclear genome. The average interval between these markers was 9.1 cM. Two genes (Mi and Mij) involved in resistance to root-knot nematodes (Meloidogyne incognita (Kofoid and White) Chitwood and Meloidogyne javanica (Treub) Chitwood) were mapped to a single linkage group (Group I). These two loci were separated by a 16.5-cM interval. One codominant AFLP marker (EAA/MCAT10) was tightly linked to the Mij locus (3.4 cM), and a dominant AFLP marker (EAT/MCAT2) was found to be closely associated with the Mi locus (6.0 cM). These markers are being studied for utilization in peach rootstock breeding with marker-assisted selection.Key words: peach rootstocks, root-knot nematodes, resistance, AFLP, mapping.