Microsatellite loci in Japanese quail and cross-species amplification in chicken and guinea fowl

In line with the Gifu University''s initiative to map the Japanese quail genome, a total of 100 Japanese quail microsatellite markers isolated in our laboratory were evaluated in a population of 20 unrelated quails randomly sampled from a colony of wild quail origin. Ninety-eight markers were polymorphic with an average of 3.7 alleles per locus and a mean heterozygosity of 0.423. To determine the utility of these markers for comparative genome mapping in Phasianidae, cross-species amplification of all the markers was tested with chicken and guinea fowl DNA. Amplification products similar in size to the orthologous loci in quail were observed in 42 loci in chicken and 20 loci in guinea fowl. Of the cross-reactive markers, 57.1% in chicken and 55.0% in guinea fowl were polymorphic when tested in 20 birds from their respective populations. Five of 15 markers that could cross-amplify Japanese quail, chicken, and guinea fowl DNA were polymorphic in all three species. Amplification of orthologous loci was confirmed by sequencing 10 loci each from chicken and guinea fowl and comparing with them the corresponding quail sequence. The microsatellite markers reported would serve as a useful resource base for genetic mapping in quail and comparative mapping in Phasianidae.     

Chicken microsatellite primers are not efficient markers for Japanese quail

Domestic fowl or chicken (Gallus gallus) and Japanese quail (Coturnix japonica) belong to the family Phasianidae. The exchange of marker information between chicken and quail is an important step towards the construction of a high-resolution comparative genetic map in Phasianidae, which includes several poultry species of agricultural importance. We tested chicken microsatellite markers to see if they would be suitable as genetic linkage markers in Japanese quail. Twenty-six per cent (31/120) of chicken primers amplified individual loci in Japanese quail and 65% (20/31) of the amplified loci were found to be polymorphic. Eleven of the polymorphic loci were excluded as uninformative because of the lack of amplification in some individuals or high frequency of nonspecific amplification. The sequence information of the remaining nine loci revealed six of them to contain microsatellites that were nearly identical with those of the orthologous regions in chicken. For these six loci, allele frequencies were estimated in 50 unrelated quails. Although the very few chicken markers that do work well in quail could be used as anchor points for a comparative mapping, most chicken markers are not useful for studies in quail. Therefore, more effort should be committed to developing quail-specific markers rather than attempting to adapt chicken markers for work in quail.     

Use of chicken microsatellite markers in turkey: a pessimistic view

Eighty-eight chicken microsatellite markers, previously developed in our laboratory, were tested for their ability to amplify polymorphic fragments using turkey genomic DNA. Amplification products were obtained for 61 chicken microsatellite markers (69.1%) whereas 27 (30.9%) did not give rise to any products, even when different polymerase chain reaction conditions were employed. From the 61 markers that gave a product, only eight showed a length polymorphism while 37 were monomorphic on the three divergent commercial turkey lines used. The remaining 16 markers yielded many unspecific bands and no specific amplification product could be obtained. Five polymorphic and eleven monomorphic products contained a detectable microsatellite repeat. Furthermore, of the markers that detected a polymorphism in turkey, the observed heterozygosity (15–50%) and allelic variation (only 2 in most cases) was very low. Therefore, on the basis of our results, we think that chicken microsatellite markers are not very useful for mapping purposes in turkey.     

微卫星标记对牙鲆有丝分裂雌核发育家系的亲子鉴定

利用18个微卫星标记,对6个家系的26尾有丝分裂雌核发育牙鲆进行亲子鉴定,PCR扩增产物经8%非变性聚丙烯酰胺凝胶电泳检测,结果表明:1个座位在母本中表现为相同的基因型,视为单态座位,其他17个座位为多态;多态座位在亲子鉴定中的累计排除概率和累计个体识别概率分别为0.9985、0.9999;根据被测个体在17个微卫星座位的基因型,最后确认26尾子代的母本,其中7尾子代在某些座位表现出与其母本不完全匹配的基因型。利用微卫星标记可确定雌核发育后代的亲子关系,从而构建牙鲆雌核发育家系系谱,对牙鲆雌核发育的深入研究具有重要意义。     

Characterization of 35 novel microsatellite DNA markers from the duck (Anas platyrhynchos) genome and cross-amplification in other birds

In order to study duck microsatellites, we constructed a library enriched for (CA)n, (CAG)n, (GCC)n and (TTTC)n. A total of 35 pairs of primers from these microsatellites were developed and used to detect polymorphisms in 31 unrelated Peking ducks. Twenty-eight loci were polymorphic and seven loci were monomorphic. A total of 117 alleles were observed from these polymorphic microsatellite markers, which ranged from 2 to 14 with an average of 4.18 per locus. The frequencies of the 117 alleles ranged from 0.02 to 0.98. The highest heterozygosity (0.97) was observed at the CAUD019 microsatellite locus and the lowest heterozygosity (0.04) at the CAUD008 locus, and 11 loci had heterozygosities greater than 0.50 (46.43%). The polymorphism information content (PIC) of 28 loci ranged from 0.04 to 0.88 with an average of 0.42. All the above markers were used to screen the polymorphism in other bird species. Two markers produced specific monomorphic products with the chicken DNA. Fourteen markers generated specific fragments with the goose DNA: 5 were polymorphic and 9 were monomorphic. But no specific product was detected with the peacock DNA. Based on sequence comparisons of the flanking sequence and repeat, we conclude that 2 chicken loci and 14 goose loci were true homologous loci of the duck loci. The microsatellite markers identified and characterized in the present study will contribute to the genetic map, quantitative traits mapping, and phylogenetic analysis in the duck and goose.     

Polymorphic microsatellites in Buff-throated partridge developed by cross-species amplification

Microsatellite loci for the buff-throated partridge (Tetraophasis szechenyii), an endemic pheasant species of China, are here described for the first time. Twenty-five microsatellite markers from chicken and Japanese quail were tested on buff-throated partridge DNA by means of cross-amplification. Twenty (80%) primers yielded specific products and polymorphisms were tested in a wild population of buff-throated partridge. Twelve (48%) proved to be polymorphic with an average of two alleles per locus. Current results of buff-throated partridge microsatellites loci could be employed in population genetic studies and on other endangered pheasant species.     

Polymorphic microsatellites developed by cross-species amplifications in common pheasant breeds

Genetic variability was analysed in two common breeds of pheasant (Phasianus colchicus L. 1758) by means of cross-species amplifications of microsatellite loci: 154 chicken, Gallus gallus and 32 turkey, Meleagris gallopavo, primers were tested for amplification of pheasant DNA. Thirty-six primers (25 specific for chicken and 11 for turkey) amplified pheasant DNA. Fifteen markers yielded specific products and were tested for polymorphism. Eight of them (55%) were polymorphic, with an average polymorphism of two alleles per locus. Specific polymerase chain reaction (PCR) products were sequenced; repeats were found in 11 of the 15 markers, although only two loci showed the same repeat and could be homologous to chicken ones.     

A genetic and cytogenetic map for the duck (Anas platyrhynchos)

A genetic linkage map for the duck (Anas platyrhynchos) was developed within a cross between two extreme Peking duck lines by linkage analysis of 155 polymorphic microsatellite markers, including 84 novel markers reported in this study. A total of 115 microsatellite markers were placed into 19 linkage groups. The sex-averaged map spans 1353.3 cM, with an average interval distance of 15.04 cM. The male map covers 1415 cM, whereas the female map covers only 1387.6 cM. All of the flanking sequences of the 155 polymorphic loci--44 monomorphic loci and a further 41 reported microsatellite loci for duck--were blasted against the chicken genomic sequence, and corresponding orthologs were found for 49. To integrate the genetic and cytogenetic map of the duck genome, 28 BAC clones were screened from a chicken BAC library using the specific PCR primers and localized to duck chromosomes by FISH, respectively. Of 28 BAC clones, 24 were detected definitely on duck chromosomes. Thus, 11 of 19 linkage groups were localized to 10 duck chromosomes. This genetic and cytogenetic map will be helpful for the mapping QTL in duck for breeding applications and for conducting genomic comparisons between chicken and duck.     

使用林鹳微卫星引物对东方白鹳基因组DNA进行交叉扩增

利用林鹳11个微卫星位点的引物对东方白鹳进行交叉扩增。经过PCR体系的优化,在11个位点中有6个得到清晰的扩增条带,其余5位点得不到确切的扩增产物。对上述6个位点的扩增产物进行克隆测序分析,发现其中4个位点上的扩增产物含有微卫星重复序列,而另外两个位点中无重复单元。通过基因分型对上述4个微卫星位点进行多态性分析后发现其中的WSμ13,WSμ17位点分别为高度多态和中度多态位点,而另外两个位点则无多态性。同时还对影响交叉扩增结果成功率及微卫星位点多态性的因素进行了分析和总结。     

Construction of a genetic linkage map of Japanese quail (Coturnix japonica) based on AFLP and microsatellite markers

The Japanese quail (Coturnix japonica) is a notably valuable egg and meat producer but has also been used as a laboratory animal. In the present study, we constructed a Japanese quail linkage map with 1735 polymorphic amplified fragment length polymorphisms markers, and nine chicken microsatellite (MS) markers, as well as sex and phenotypes of two genetic diseases; a muscular disorder (LWC) and neurofilament-deficient mutant (Quv). Linkage analysis revealed 578 independent loci. The resulting linkage map contained 44 multipoint linkage groups covering 2597.8 cM and an additional 218.2 cM was contained in 21 two-point linkage groups. The total map was 2816 cM in length with an average marker interval of 5.5 cM. The Quv locus was located on linkage group 5, but linkage was not found between the LWC locus and any of the markers. Comparative mapping with chicken using orthologous markers revealed chromosomal assignments of the quail linkage group 1 to chicken chromosome 2 (GGA2), 5 to GGA22, 2 to GGA5, 8 to GGA7, 27 to GGA11, 29 to GGA1 and 45 to GGA4.     

Comparative analysis of microsatellite loci in chicken and turkey.

Cross-species amplification of 520 chicken microsatellite markers was tested by polymerase chain reaction with genomic DNA of the turkey (Meleagris gallopavo). Each primer pair was tested at six different combinations of annealing temperature and MgCl2 concentration. A total of 280 (54%) of the primer pairs produced amplification products. The majority of these products were similar, if not identical in size to those expected based on the fragment sizes of the corresponding chicken loci. Structure of the dinucleotide repeat and flanking sequences was examined for 13 turkey fragments (amplified with chicken primers) and 5 chicken fragments (amplified with turkey primers). Sequence analysis found a wide array of mutations between species in addition to differences in repeat length. To estimate the usefulness of the amplified loci for genetic mapping in the turkey, allelic polymorphism was determined for 57 of the 280 amplified loci. A total of 20 of 57 markers (35%) were polymorphic with an average of 1.4 alleles per locus. The results of this study suggest that approximately 20% of the chicken microsatellite markers will be useful for mapping the turkey genome.     

Mapping of plumage colour and blood protein loci on the microsatellite linkage map of the Japanese quail

The objective of this work was to map classical markers (plumage colours and blood proteins) on the microsatellite linkage map of the Japanese quail (Coturnix japonica). The segregation data on two plumage colours and three blood proteins were obtained from 25 three-generation families (193 F2 birds). Linkage analysis was carried out for these five classical markers and 80 microsatellite markers. A total of 15 linkage groups that included the five classical loci and 69 of the 80 microsatellite markers were constructed. Using the BLAST homology search against the chicken genome sequence, three quail linkage groups, QL8, QL10 and QL13, were suggested to be homologous to chicken chromosomes GGA9, GGA20 and GGA24, respectively. Two plumage colour loci, black at hatch (Bh) and yellow (Y), and the three blood protein loci, transferrin (Tf), haemoglobin (Hb-1) and prealbumin-1 (Pa-1), were assigned to CJA01, QL10, QL8, CJA14 and QL13, respectively.     

Fifty microsatellite markers for Japanese quail

A Japanese quail genomic library enriched for (CA/GT)n simple sequence repeats was screened and positive clones were sequenced. Fifty original microsatellite sequences were isolated that consisted mainly of perfect repeats of the dinucleotide (CA/GT)n motif and a corresponding number of polymerase chain reaction (PCR) primer pairs complementary to unique DNA sequences flanking the microsatellite repeats were designed to detect the repeats. Forty-six percent (23 of 50) of the markers revealed polymorphism in two unrelated quail individuals (one male and one female) randomly sampled from a population of wild quail origin. All 50 primer pairs were tested in the PCR for their ability to amplify chicken genomic DNA. Amplification products were obtained for 14 (28.0%) of the markers at the annealing temperature optimized for quail. These results provide an opportunity to begin characterizing the quail genome for the development of a genetic map for this economically valuable species and the eventual construction of a comparative genetic map in Phasianidae, which comprises a number of agriculturally important species of poultry.     

Development and multiplex PCR amplification of novel microsatellite markers in the White‐tailed Sea Eagle,Haliaeetus albicilla (Aves: Falconiformes,Accipitridae)

We report the development of 14 novel polymorphic microsatellite markers cloned from the White‐tailed Sea Eagle, Haliaeetus albicilla, a formerly threatened raptor that has received much conservation attention throughout Eurasia. We also present a protocol for multiplex polymerase chain reaction (PCR) amplification of the loci. Among 40 unrelated H. albicilla individuals from southern Sweden, the markers produced two to eight alleles per locus, and average observed and expected heterozygosities were 0.463 and 0.468, respectively. We further present five microsatellite markers that appeared monomorphic in H. albicilla, but which may be of interest for use in other raptor species.     

Cross‐species amplification of zebrafish and central stoneroller microsatellite loci in six other cyprinids

One hundred and twenty Danio rerio and Campostoma anomalum microsatellite loci were tested for amplification in the cyprinids Phoxinus phoxinus, Scardinius erythrophthalmus, Rutilus rutilus, Abramis brama, Carassius carassius and Gobio gobio . The PCR amplification products were recorded in 35 to 50 loci, depending on focal species. Eight to 28 of the amplifying loci were also polymorphic, establishing potential sets of genetic markers for population genetics studies.     

Isolation and characterization of microsatellites in Pachygrapsus marmoratus (Grapsidae; Decapoda; Brachyura)

Seven polymorphic microsatellite loci were isolated and characterized from the grapsid crab Pachygrapsus marmoratus using FIASCO (fast isolation by AFLP of sequences containing repeats) protocol. Twenty‐seven primer pairs were designed, 14 of which worked well in polymerase chain reaction (PCR), amplifying a fragment of the expected size. Variability was tested in 18 specimens collected along the coast of Tuscany (Italy). Five loci were discarded due to stutter products during their amplification, and two resulted to be monomorphic. The remaining seven loci, showed a number of alleles ranging from two to 14 and an observed heterozygosity ranging from 0.12 to 0.67.     

Allelic variation and genetic linkage of avian microsatellites in a new turkey population for genetic mapping

Efforts to build a comprehensive genetic linkage map for the turkey (Meleagris gallopavo) have focused on development of genetic markers and experimental resource families. In this study, PCR amplification was attempted for 772 microsatellite markers that had been previously developed for three avian species (chicken, quail and turkey). Allelic polymorphism at 410 markers (53.1% of total examined) was determined by genotyping ten individuals (six F1 parents and four grandparents) in a new resource population specifically developed for genetic linkage mapping. Of these 410 markers, 109 (26.6%) were polymorphic in the tested individuals, with an average of 2.3 alleles per marker. Higher levels of polymorphism were found for the turkey-specific markers (61.1%) than for the chicken (22.7%) or quail-specific markers (33.3%). To test the fidelity of the matings, demonstrate the power of these families for linkage analysis, and determine genetic linkage relationships, 86 polymorphic markers were genotyped for up to 224 birds including founder grandparents, parents and F2 progeny. Linkage relationships for many of the chicken markers elucidated in the turkey were comparable to those observed in the chicken. These data demonstrate that the new UMN/NTBF resource population will provide a solid foundation for constructing a comparative genetic map of the turkey.     

A panel of microsatellite loci for population studies of Sakhalin taimen Parahucho perryi (Brevoort)

Using DNA samples of Sakhalin taimen and a set of microsatellite loci, earlier reported for other salmonid fishes (Salmonidae), successful cross-species amplification was performed. A total of 56 Sakhalin taimen (Parahucho perryi) samples from the Daga, Nabil, Poronai, and Agnevo rivers (Sakhalin Island) were examined at 36 microsatellite loci, most of which were described for other species and first tested in taimen. Among the 21 loci first tested in taimen, two loci produced no amplification products. The remaining 19 loci were successfully amplified (for some loci, new primers were generated). Thirteen of these loci were monomorphic, while six loci were polymorphic and used in further population genetic analysis. In addition, with the purpose of modification of the allele sizes and optimization of the research technique, new primers for the already known 12 loci of Sakhalin taimen were designed. Three more loci were included in analysis without changes. As a result, a joint panel consisting of 21 polymorphic microsatellite markers was suggested for analysis of Sakhalin taimen. This panel was tested with four population samples from the rivers of Sakhalin Island.The results showed that this panel of markers could be used in detailed population studies for evaluation of the level of genetic differentiation, inbreeding, and migrations in Sakhalin taimen, an endangered species with a fragmented range. Using this approach in further studies will make it possible to isolate basic populations, which is necessary for conservation of this rare species.     

A preliminary microsatellite genetic map of the ostrich (Struthio camelus)

Molecular genetic maps can provide information for the identification and localization of major genes associated with quantitative traits. However, there are currently no published genetic linkage maps for any ratites. Herein, a preliminary genetic map of ostrich was developed using a two-generation ostrich reference family by linkage analysis of 104 polymorphic microsatellite markers, including 40 novel markers reported in this study. A total of 35 microsatellite markers were placed into 13 linkage groups. Five linkage groups are composed of three or more loci, whereas the remaining eight groups each contained two markers. The sex-averaged map spans 365.4 cM. The marker interval of each linkage group ranges from 5.3 to 25.4 cM, and the average interval distance is 16.61 cM. The male map covers 342.7 cM, with an average intermarker distance of 15.58 cM, whereas the female map is 456.7 cM, with the average intermarker spacing of 20.76 cM. In order to screen the orthologous loci between ostrich and chicken, all of the flanking sequences of the 104 polymorphic loci, nine monomorphic loci and a further 12 reported microsatellite loci for ostrich were screened against the chicken genomic sequence using the BLAST algorithm (Altschul et al., 1990), and corresponding orthologs were found for 13 sequences. The microsatellite loci and genetic map developed in this study will be useful for QTL mapping, population genetics and phylogenetic studies in the ratite. In addition, the 13 orthologous loci identified in this study will be advantageous to the construction of a comparative genetic map between chicken and ostrich.     

Characterization of polymorphic microsatellite markers for the Carnaby's cockatoo (Calyptorhynchus latirostris) and related black cockatoo species

Microsatellite loci were isolated from Carnaby's black cockatoo (Calyptorhynchus latirostris: Aves), a highly valued, endangered, and endemic species of bird from Western Australia. This study describes three dinucleotide and one tetranucleotide microsatellite loci for which the primers produced clear and polymorphic amplification patterns with between two and nine alleles and moderate levels of variability. Two additional dinucleotide markers which were monomorphic in the Carnaby's cockatoo were able to amplify and were polymorphic in two other species of black cockatoo, greatly increasing the utility of these markers.