Patterns of gene flow and population genetic structure in the canyon treefrog, Hyla arenicolor (Cope)

Patterns of gene flow and genetic structuring were examined in the canyon treefrog, Hyla arenicolor (Cope). Hierarchical analysis of genetic variation was performed on mitochondrial cytochrome b haplotypes from 323 individuals, representing 32 populations from previously described phylogeographic regions. Results from AMOVA revealed that 60.4-78.9% of the recovered genetic variation was the result of differences in the appointment of genetic variation between subdivisions of the primary phylogeographic regions. In contrast, populations only contained between 13.9 and 30.1% of the observed haplotypic variation. Gene flow estimates based on calculations of phi ST revealed moderate levels of gene flow within phylogeographic regions, but there was no evidence of gene flow between these regions, suggesting that geographical boundaries were probably important in the formation of phylogeographic structure in H. arenicolor. Phylogeographic regions exhibited very different patterns of gene flow. One region showed evidence of recent colonization. Another region exhibited very limited gene flow. Moderate to high estimates of gene flow were obtained for populations from two distinct phylogeographic regions characterized by mesic and xeric environments. Isolation by distance was observed in both regions suggesting that these regions are in genetic equilibrium. Because gene flow is extremely unlikely between the populations in the xeric region, this result is interpreted as historical gene flow. These results indicate that isolation-by-distance effects may still be observed even when population genetic structure and gene flow are the result of historical association.     

Phylogeography of the canyon treefrog, Hyla arenicolor (Cope) based on mitochondrial DNA sequence data

Patterns of phylogeography and gene flow were examined in the canyon treefrog, Hyla arenicolor. A total of 973 bp of mitochondrial cytochrome b sequence data were obtained for 65 individuals from 53 populations, yielding 50 unique haplotypes. Interpopulation sequence variation ranged from 0 to 13.7%. Phylogenetic analysis revealed three deeply divergent mtDNA lineages. These three Clades were mapped onto geography and found to represent completely concordant, nonoverlapping, geographical regions. Levels of sequence divergence between the three Clades were equal to or greater than published levels of divergence found in other vertebrate species and genera. Furthermore, one Clade of H. arenicolor was found to be more closely related to the outgroup H. eximia than to other H. arenicolor, suggesting that the taxonomy of this species may require revision.     

Geographical variation in male advertisement calls and female preference of the wide‐ranging canyon treefrog,Hyla arenicolor

We surveyed the geographical variation in male advertisement calls of the wide‐ranging canyon treefrog, Hyla arenicolor, and found large call differences among geographically distant lineages that had been characterized by a recent phylogeographical study. To test whether these call differences were biologically relevant and could allow reproductive isolation of different lineages should they come into secondary contact, we assessed female preference in a lineage occurring in southern Utah and north‐western Arizona, USA. These females exhibited a strong preference for their own lineage's call type over the calls of two Mexican lineages, but not over the calls from the geographically nearest lineage. We also identified traits that female frogs probably use to discriminate between lineage‐specific advertisement calls. Our behavioural results, together with recent molecular estimates of phylogenetic relationships among lineages, will guide future work addressing the evolutionary forces that have led to this biologically significant variation in male sexual signals. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ??, ??–??.     

Suitability of AFLP markers for the study of genetic relationships among Korean native dogs.

To determine the genetic relationships among domestic dog breeds, we performed both a sequence comparison of mitochondrial DNA (mtDNA) and an amplified fragment length polymorphisms (AFLP) analysis. Three of four regions of mtDNA, cytochrome b, cytochrome oxidase subunit II, and 16S rRNA genes were highly homogeneous among dog breeds, whereas the other region, the control region, showed relatively high polymorphisms with a maximum percentage difference of 3.18%. However, the control region showed extensive polymorphism even within breeds, and the relationship tree derived from the data could not clearly delimit distinct breeds. 19 EcoRI/MseI primer combinations were used to generate AFLP markers among 25 dogs from 11 breeds including three Korean native dogs. These amplification reactions allowed the detection of more than 1900 amplification products of which 408 were identified as polymorphic bands. Unrooted neighbor-joining tree based on dissimilarity values showed that the Korean native dogs were clustered together with the Asian dogs and that the Asian originated dogs were clustered separately from Western originated dogs. A consensus tree using parsimony method also showed Korean native dogs were grouped separately from the other dogs with moderate bootstrap values. Taken together, it is concluded that AFLP analysis is a more informative tool for revealing genetic relationships among dog breeds than mtDNA sequence comparison.     

Molecular patterns of differentiation in canyon treefrogs (Hyla arenicolor): evidence for introgressive hybridization with the Arizona treefrog (H. wrightorum) and correlations with advertisement call differences

Detection of genetic and behavioural diversity within morphologically similar species has led to the discovery of cryptic species complexes. We tested the hypothesis that US populations of the canyon treefrog (Hyla arenicolor) may consist of cryptic species by examining mate‐attraction signals among three divergent clades defined by mtDNA. Using a multi‐locus approach, we re‐analysed phylogenetic relationships among the three clades and a closely related, but morphologically and behaviourally dissimilar species, the Arizona treefrog (H. wrightorum). We found evidence for introgression of H. wrightorum’s mitochondrial genome into H. arenicolor. Additionally, the two‐clade topology based on nuclear data is more congruent with patterns of call variation than the three‐clade topology from the mitochondrial dataset. The magnitude of the call divergence is probably insufficient to promote isolation of the nuclear DNA‐defined clades should they become sympatric, but further divergence in call properties significant in species identification could promote speciation in the future.     

Analysis of genetic relationships among Rosa damascena plants grown in Turkey by using AFLP and microsatellite markers

Rosa damascena Mill. is the most important rose species for rose oil production. The main rose oil producers in the world are Turkey and Bulgaria and they obtain the rose oil almost exclusively from R. damascena. In spite of coming from the same original populations, R. damascena plants grown in Turkey show some morphological differences. In this study, it was aimed to investigate the genetic relationships among R. damascena plants grown in Turkey by using microsatellite and AFLP markers. Twenty three AFLP and nine microsatellite primer pairs were used for this aim. No polymorphism could be detected among the plants, as the marker patterns obtained from different plants are identical. The conclusion from these data is that all R. damascena plants under study are derived from the same original genotype by vegetative propagation. Furthermore, the observed morphological differences originate from point mutations not detectable by molecular markers. Therefore, they are equivalent to sport mutations frequently observed in cut and garden rose varieties.     

Analysis of genetic diversity and sectional relationships in Musa using AFLP markers

The AFLP technique was used to assess the genetic diversity and sectional relationships in 39 accessions representing the four main sections of the genus Musa. Eight AFLP + 3 primer pairs produced 260 polymorphic bands that were used in cluster and PCO analysis. A wide range of variability was observed among the species within the sections of the genus Musa. AFLP data was useful in separating the different sections of the genus as well as differentiating the different genomic groups of section Eumusa. Section Rhodochlamys (x = 11) appeared as a distinct entity and clustered closely with the Musa acuminata Colla complex of section Eumusa that has the same basic chromosome number. This relationship is congruent with previous studies. However, unlike previous proposals that questioned the identity of Rhodochlamys as a separate taxonomic unit, PCO analysis of the AFLP data showed that it is a distinct entity. Musa laterita Cheesman (Rhodochlamys) and Musa schizocarpa Simmonds clustered with the M. acuminata complex suggesting that they may be sources of useful genes for the improvement of the cultivated bananas. Callimusa formed a distinct unit and was closer to Australimusa than to the other sections. Although both sections share the same basic chromosome number of x = 10 these sections are genetically distinct     

Assessment of genetic diversity and interspecific relationships among three species of Podophyllum using AFLP markers and podophyllotoxin content

The genus Podophyllum (common name: May Apple) has high medicinal value due to the presence of anticancer molecule, podophyllotoxin. A total of 35 individuals belonging to three species of Podophyllum viz. P. hexandrum Royle, P. sikkimensis R. Chatterjee and Mukherjee both Indian species, and their American counterpart, P. peltatum L. have been investigated with a view to ascertain variation in their (1) podophyllotoxin content, and (2) molecular profiles generated through AFLP markers. The active principle content varied within the representative individuals of different populations of a species and between species; the species-wise podophyllotoxin content (% of dry wt) ranged as follows: P. hexandrum-Munsyari populations: 0.39–1.20 %, P. hexandrum-Kullu populations: 0.58–1.50 % (highest), P. peltatum: 0.50–1.30 %, and P. sikkimensis: 0.06–0.73 % (lowest). Detection of podophyllotoxin in P. sikkimensis, although at low levels, would appear to be the first report of its occurrence in this species. The genetic diversity and relationship amongst 35 sampled individuals of three species have been analyzed using 20 AFLP markers, which resulted into 1,358 loci of which 595 were polymorphic revealing 44 % polymorphism. High level of genetic diversity was observed (percent of polymorphic bands, PPB = 88.01 %; PIC = 0.813) among the species, while it was low within the individual species (PIC = 0.57 %; Marker index, MI = 4.77). Genetic similarity among the species (calculated with Euclidean coefficient) showed two major clusters. Cluster one contained all the individuals of P. peltatum (American May Apple) whereas cluster two grouped together individuals representing various populations, belonging to both the species of Indian May Apple (P. hexandrum, and P. sikkimensis). The observed paired relationship (45–50 % similarity; calculated from AFLP data) of intercontinental species in the Podophyllum group (P. hexandrum, and P. sikkimensis vs. P. peltatum) would appear to be paraphyletic. The AFLP data of the analyzed representatives have been used to examine the sister relationships among these species, and would be beneficial to find ways to strengthen the gene flow among populations to maintain the natural genetic variation within the populations of Podophyllum species.     

Genetic relationships among Aster species by multivariate analysis and AFLP markers

Fourteen species of Aster were characterized for morphological traits of commercial interest, chromosome number and genetic diversity by AFLP markers to exploit this material not only for breeding purposes but also as source of pharmaceuticals. The variation observed among the Aster species for five morphological traits was summarized by means of discriminant analysis. Three significant canonical variables, accounting for about 96% of total variance, were mainly correlated with capitulum diameter, number of ligulae and leaf length. The morphological variation appeared related to ploidy level of the species. A diploid chromosome number ranging from 10 to 18 was evident in seven species whereas the other species are polyploids with variable chromosome numbers up to 52. The pollen production and stainability were quite variable among Aster species. Furthermore, seven species produced big pollen grains besides normally-sized ones, at a frequency ranging from 1 to 12%, possibly due to 2n pollen production. The possibility that evolution of Aster genus could be driven by 2n gametes and sexual polyploidization is discussed. Implications of 2n gametes for Aster breeding are also mentioned. AFLP analysis of the fourteen Aster species based on six primer combinations revealed a total number of 421 polymorphic amplified DNA fragments. The genetic similarities estimated between the Aster species, based on both shared and unique amplification products ranged from 0.335 to 0.604 across all species and revealed a geographically based clustering within the genus. The highest similarity was detected between A. alpinus and A. amellus with Eurasian origin.     

Geographical diversity and genetic relationships among Cedrus species estimated by AFLP

Genetic diversity was described in 17 cedar populations covering the geographical range of the four species of the genus Cedrus. The study was conducted using amplified fragment length polymorphism (AFLP) on haploid tissues (megagametophytes). Eleven selective AFLP primer pairs generated a total of 107 polymorphic amplification products. Correspondence and genetic distance analyses indicated that Cedrus deodara constitutes a separate gene pool from the Mediterranean cedars. Within Mediterranean cedars, we distinguished two groups: the first one is made of Cedrus atlantica, while the second one is made of Cedrus libani and Cedrus brevifolia, these latter two species being genetically similar despite important divergence previously observed for morphological and physiological traits. The lowest intrapopulation variability was found in the two C. deodara populations analyzed. Surprisingly, C. brevifolia, the endemic taxon from the island of Cyprus that is found in small and fragmented populations, showed one of the highest levels of diversity. This unexpected pattern of diversity and differentiation observed for C. brevifolia suggests a recent divergence rather than a relictual, declining population. Patterns of diversity within- and among-populations were used to test divergence and fragmentation hypotheses and to draw conclusions for the conservation of Cedrus gene pools.     

Genetic relationships among Mediterranean Pistacia species evaluated by RAPD and AFLP markers

Polymorphisms among Mediterranean basin Pistacia species and accessions within species were assessed by random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) analyses. Twenty-eight Pistacia accessions representing six species from geographically diverse locations in the Mediterranean area were analyzed. With RAPD, a total of 259 DNA fragments were amplified by 27 pre-selected primers, 254 were polymorphic fragments. AFLP analysis with 15 primer sets, produced 954 (93%) polymorphic bands out of a total of 1026. A Mantel test revealed an extremely high correlation (r=0.99) between similarity matrices generated from RAPD and AFLP data sets, indicating that similar results were obtained by the two techniques. Dendrograms constructed from the similarity matrices showed that Pistacia species could be clustered into two groups, one group containing all the #E5/E5#. lentiscus and the second group containing all other accessions. The latter group was divided into two subgroups, one consisting of #E5/E5#. palaestina and #E5/E5#. terebinthus; the other consisting of #E5/E5#. atlantica, #E5/E5#. khinjuk and #E5/E5#. vera. P. vera and P. khinjuk were highly similar, as were P. palaestina and P. terebinthus.     

Frequency selectivity of hearing in the green treefrog,Hyla cinerea

Frequency selectivity of hearing was measured in the green treefrog, Hyla cinerea. A psychophysical technique based on reflex modification was used to obtain masked threshold estimates for pure tones (300-5,400 Hz) presented against two levels of broadband masking noise. A pure tone (S-1) presented 200 ms prior to a reflex-eliciting stimulus (S-2) inhibited the motor reflex response to S-2. The magnitude of this reflex modification effect varied systematically with the sound pressure level (SPL) of S-1, and threshold was defined as the SPL of S-1 at which the reflex modification effect disappeared. Masked thresholds were used to calculate critical ratios, an index of the auditory system's frequency selectivity. The frequency selectivity of the treefrog's hearing is greatest and critical ratios are lowest (22-24 dB) at about 900 and 3,000 Hz, the two spectral regions dominant in the male treefrog's species-specific advertisement call. These results suggest that the treefrog's auditory system may be specialized to reject noise at biologically-relevant frequencies. As in other vertebrates, critical ratios remain constant when background noise level is varied; however, the shape of the treefrog's critical ratio function across frequencies differs from the typical vertebrate function that increases with increasing frequency at a slope of about 3 dB/octave. Instead, the treefrog's critical ratio function resembles its pure tone audiogram. Although the shape of the treefrog's critical ratio function is atypical, the critical ratio values themselves are comparable to those of many other vertebrates in the same frequency range. Critical ratio values here measured behaviorally do not match critical ratio values previously measured physiologically in single eighth nerve fibers.(ABSTRACT TRUNCATED AT 250 WORDS)     

Association analysis,genetic diversity and structure analysis of tobacco based on AFLP markers

Knowledge in the area of genetic diversity could aid in providing useful information in the selection of material for breeding such as hybridization programs and quantitative trait loci mapping. To this end, 50 Nicotiana tabacum genotypes were genotyped with 21 primer combination of amplified fragment length polymorphism (AFLP). A total of 480 unambiguous DNA fragments and 373 polymorphic bands were produced with an average of 17.76 per primer combination. Also, the results revealed high polymorphic rate varing from 52.63 to 92.59 %, demonstrating that AFLP technique utilized in this research can be a powerful and valuable tool in the breeding program of N. tabacum. Cluster analysis based on complete linkage method using Jaccard’s genetic distance, grouped the 50 tobacco genotypes into eight clusters including three relatively big clusters, one cluster including Golden gift, Burly 7022 and Burly Kreuzung, one cluster consisting of two individuals (Pereg234, R9) and three single-member clusters (Pennbel69, Coker176 and Budisher Burley E), Recent genotypes showed high genetic distance from other genotypes belonging to cluster I and II. Association analysis between seven important traits and AFLP markers were performed using four statistical models. The results revealed the model containing both the factors, population structure (Q) and general similarity in genetic background arising from shared kinship (K), reduces false positive associations between markers and phenotype. According to the results nine markers were determined that could be considered to be the most interesting candidates for further studies.     

Using AFLP markers and the Geneland program for the inference of population genetic structure

The use of dominant markers such as amplified fragment length polymorphism (AFLP) for population genetics analyses is often impeded by the lack of appropriate computer programs and rarely motivated by objective considerations. The point of the present note is twofold: (i) we describe how the computer program Geneland designed to infer population structure has been adapted to deal with dominant markers; and (ii) we use Geneland for numerical comparison of dominant and codominant markers to perform clustering. AFLP markers lead to less accurate results than bi-allelic codominant markers such as single nucleotide polymorphisms (SNP) markers but this difference becomes negligible for data sets of common size (number of individuals n≥100, number of markers L≥200). The latest Geneland version (3.2.1) handling dominant markers is freely available as an R package with a fully clickable graphical interface. Installation instructions and documentation can be found on http://www2.imm.dtu.dk/~gigu/Geneland.     

Use of AFLP and RAPD molecular genetic markers and cytogenetic analysis to explore relationships among taxa of the Patagonian Bromus setifolius complex

Bromus setifolius var. pictus (Hook) Skottsb., B. setifolius var. setifolius Presl. and B.setifolius var. brevifolius Ness are three native Patagonian taxa in the section Pnigma Dumort of the genus Bromus L. AFLP and RAPD analysis, in conjunction with genetic distance measurements and statistical techniques, revealed variation within this group and indicated that B. setifolius var. brevifolius was closely related to B. setifolius var. pictus, with both taxa being more distantly related to B. setifolius var. setifolius. Cytogenetic analysis confirmed the chromosomal number of B. setifolius var. pictus (2n = 70) and B. setifolius var. setifolius (2n = 28) and showed for the first time that B. setifolius var. brevifolius had 2n = 70. The combination of molecular genetic and cytogenetic evidence supported a species status for two of the three taxa and suggested hypotheses for the evolutionary origin of these complex taxa. Species status was also indicated for B. setifolius var. setifolius. Based on these findings, we suggest that B. setifolius var. pictus be referred to as B. pictus Hook var. pictus, and B. setifolius var brevifolius as B. pictus Hook var brevifolius. The correlation between AFLP diversity and variation in ecological parameters suggested that this marker system could be used to assess breeding progress and to monitor the domestication of Patagonian Bromus species for agronomic use.     

广西毛竹种质资源AFLP分析

采用扩增片段长度多态性分子标记方法,对收集于广西的11个种源33份毛竹种质进行了分析.结果表明：5对引物组合用于选择性扩增,共得到198条扩增带,其中多态性带47条(23．74％);11个毛竹种源间的亲缘关系较近,但仍存在着一定的遗传变异,它们之间的遗传距离 D 在0．0006~0．1369之间;UPGMA 聚类分析,可将供试的11个毛竹种源分为4大类,其中昭平种源和南丹种源各为一类,与其它两类较易区分. PCA 分析结果与聚类分析结果相一致;Mantel 检验结果表明毛竹各种源遗传距离与地理距离间相关性显著(r ＝0．5558,P ＝0．0160).说明 AFLP 分子标记能将 11个种源33份毛竹种质区分开,是毛竹种质资源鉴别的有效手段和方法.