Effect of salicylic acid on protein composition of Tatar buckwheat Fagopyrum tataricum calluses with different ability for morphogenesis

The effect of salicylic acid on the content of soluble proteins and individual polypeptides in Tatar buckwheat Fagopyrum tataricum calluses differing in ability for morphogenesis was studied. Changes in the protein composition of the calluses cultivated in the dark and in the light indicated the higher sensitivity of the non-morphogenic callus. Different response of callus cultures to salicylic acid and conditions of cultivation (light, darkness) is suggested to be associated with the antioxidant defense system, which is, in particular, characterized by the hydrogen peroxide content in the calluses. Salicylic acid increased the H₂O₂ content in non-morphogenic calluses more strongly than in morphogenic calluses, and the difference was more significant for the calluses cultivated in the light.Translated from Biokhimiya, Vol. 70, No. 3, 2005, pp. 390–396.Original Russian Text Copyright © 2005 by Maksyutova, Galeeva, Rumyantseva, Viktorova.     

Development and characterization of ten novel microsatellite markers from olive ridley sea turtle (Lepidochelys olivacea)

Olive ridleys, one of the widely distributed marine turtle species has undergone declines in recent years due to multiple anthropogenic factors warranting conservation efforts for which assessment of genetic variability in existing populations become critical. Here we describe development of ten new microsatellite markers from a short sequence repeat-enriched partial genomic DNA library, which are found to be highly informative for genetic studies. Eight of these markers when tested on 83 olive ridley turtles revealed high allelic diversity (4–27 alleles per marker), and high observed and expected heterozygosity estimates that ranged from 0.29 to 0.82 and 0.62 to 0.94, respectively. Two microsatellites were monomorphic in the tested olive ridley samples, but were found to be informative/polymorphic when tested on related marine turtle species. More importantly, nine of the new markers showed robust cross-species amplifications in three related species Dermochelys coriacea, Chelonia mydas and Eretmochelys imbricata. Thus, this study describes ten new microsatellite markers and also demonstrates their potential as efficient genetic markers in studies related to parentage analysis, population structure, phylogeography and species relationships of olive ridleys and other marine turtle species.     

Development of microsatellite markers in the marine phytoplankton Karenia mikimotoi (Dinophyceae)

The marine phytoplankton, Karenia mikimotoi, causes severe red tides which are associated with mass mortality of marine fish, and have expanded their distributions in the coastal waters of western Japan. To assess the dispersal mechanism, a population genetic study using highly polymorphic genetic markers is one of the crucial approaches. Here we developed 12 polymorphic microsatellite markers from K. mikimotoi. These loci provide a class of highly variable genetic markers, as the number of alleles ranged from 5 to 23, and the estimate of gene diversity was from 0.551 to 0.933 across the 12 microsatellites. We consider these loci potentially useful for detailing the genetic structure and gene flow among K. mikimotoi populations.     

Development of twenty-five polymorphic microsatellite markers for the endangered red-cockaded woodpecker (Picoides borealis)

Twenty-five polymorphic microsatellite markers were developed for the endangered red-cockaded woodpecker (Picoides borealis). The number of alleles ranged from two to five and observed heterozygosities ranged from 0.036 to 0.750. These loci should be useful tools for conducting research towards the management and conservation of this species.     

Isolation of polymorphic microsatellite markers from <Emphasis Type="Italic">Przewalskia tangutica</Emphasis> (Solanaceae)

Microsatellite-containing regions were isolated and characterized in Przewalskia tangutica Maxim (Solanaceae), an endemic and endangered species to the Qinghai-Tibetan Plateau of China. An enrichment protocol yielded 200 positive clones. We designed primers to amplify 29 unique microsatellites, 12 of which amplified cleanly and were polymorphic. A survey of 17 individuals showed that these loci are highly variable with the number of alleles ranging from 3 to 12, and expected heterozygosity ranged from 0.2929 to 0.4947. Those markers will be useful for studies of population structure and intraspecific variation in P. tangutica.     

Development and application of microsatellite markers for genomic analysis of papaya

Papaya has a relatively small genome, displays high levels of phenotypic diversity, and is amenable to transformation, making it attractive as a fruit tree model system. The high level of phenotypic diversity seen among papaya cultivars in the field does not correlate with the low levels of genotypic polymorphism thus far elucidated. The highly mutable nature of microsatellites or simple sequence repeats (SSRs) make them potentially powerful markers for distinguishing deoxyribonucleic acid (DNA) polymorphisms between closely related genotypes. Genomic research for papaya has resulted in a significant quantity of sequence data. We mined 28.1 Mb of bacterial artificial chromosomes end sequences, 5.8 Mb of complementary DNA, and 1.6 Mb of random genomic sequences for SSRs. We generated 938 SSR markers and tested for polymorphism among seven varieties that had been used to produce five mapping populations. The level of polymorphism was highest for Kaek Dum × 2H94 with 210 markers, followed by UH928 × SunUp with 194, AU9 × SunUp with 189, UH918 × SunUp with 177, and Kapoho × SunUp displaying the lowest level with 97. Variation in levels of polymorphism, motif predominance, and motif length between the genomic and genic fractions indicated differential selection pressures acting on the microsatellites in these two fractions. The microsatellites developed in this study will greatly assist in the genetic and physical mapping of the papaya genome as well as enhance breeders’ ability to improve the crop. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Development and characterization of microsatellite loci for lotus (Nelumbo nucifera)

This paper reports the development of microsatellite primers for Nelumbo nucifera Gaerten. By screening genomic libraries enriched with 10 kinds of probes, Seventeen polymorphic loci were isolated and primers were designed. Polymorphism of these 17 loci was assessed in 24 individuals. All the 17 loci are polymorphic and the number of alleles ranged from two to seven. Observed heterozygosity and expected heterozygosity ranged from 0.0000 to 0.9176 and from 0.2837 to 0.7917 respectively. These microsatellite loci should be useful for studying the genetic diversity of N. nucifera.     

Development of 10 polymorphic microsatellite loci primers for Codonopsis pilosula Nannf. (Campanulaceae)

Codonopsis pilosula Nannf., as an important traditional Chinese medicinal plant species, has been used to treat fatigue, thirst and loss of appetite. In this study, we developed 10 new microsatellite loci primers from the genome of this species using the combined biotin capture method. The polymorphism of each locus was further assessed through 27 individuals from four geographically distant populations. The number of alleles per locus ranged from 6 to 11. The observed and expected heterozygosity ranged from 0.15 to 0.24 and 0.27 to 0.40, respectively. We further performed cross-priming tests of these loci in the other three congeneric species. These microsatellite markers will be useful for investigating genetic diversity within and between populations of these species.     

Development and characterization of new microsatellite markers for ginseng (<Emphasis Type="Italic">Panax ginseng</Emphasis> C. A. Meyer)

Panax ginseng C.A. Meyer, commonly known as Korean or Asian ginseng, is a perennial herb native to Korea and China. Its roots are highly prized for several medicinal properties. The present study describes development and characterization of twenty-two polymorphic microsatellite markers for this species. A total of 99 alleles were detected with an average of 4.5 alleles per locus across 20 accessions. Values for observed (H _O ) and expected (H _E ) heterozygosities ranged from 0.05 to 1.00 and from 0.18 to 0.73, respectively. Eleven loci deviated from Hardy–Weinberg equilibrium (P < 0.001). Significant (P < 0.05) heterozygote deficiency was observed at 13 loci. Exact test for linkage disequilibrium showed significant values (P < 0.05) between 12 pairs of loci. These microsatellite markers provide powerful tools for understanding population and conservation genetics of this species and also for genetic differentiation and authentication of different Panax species being used in commercial ginseng products.     

基于转录组平台的蛤仔微卫星标记筛选

以菲律宾蛤仔转录组测序所得拼接序列为基础,采用MISA软件进行微卫星分析,对其中的145个微卫星位点进行引物设计,得到具有清晰扩增条带的微卫星位点58个。对大连庄河野生蛤仔群体的扩增结果表明,18个位点显示单态性,40个位点表现为多态性。该群体40个多态性微卫星位点得到的等位基因数在2—6之间,平均等位基因数为3.4250±0.9718,观测杂合度和期望杂合度分别在0.0000—1.0000和0.0615—0.7996之间,平均值分别为0.2727±0.2272和0.4739±0.1902,群体平均Nei指数为0.4664±0.1872。多态信息含量(PIC)在0.0586—0.7529之间,平均值为0.4148±0.1707,其中16个微卫星位点的PIC值大于0.5,为高度多态性,15个位点0.25PIC0.5,为中度多态性,其余9个为低度多态性。经Sequential Bonferroni校正的Hardy-Weinberg平衡检验,有10个位点尚未偏离平衡。基于转录组平台筛选微卫星标记的方法,在很大程度上推动了DNA分子标记的开发。研究开发的微卫星标记可用于蛤仔群体遗传学、遗传连锁图谱构建及其他相关研究,为蛤仔分子标记辅助育种及群体种质保护等工作提供技术支持。     

Development of microsatellite markers for noninvasive DNA samples of the eastern green lizard Lacerta viridis

We report the isolation and characterization of microsatellite markers for the eastern green lizard (Lacerta viridis) usable for noninvasive samples. Ten polymorphic loci were obtained by screening 3000 recombinant clones and tested on 39 noninvasive DNA samples of individuals from different locations of the Danube area, Germany. Allelic richness ranged from three to nine alleles, the observed heterozygosity from 0.33 to 0.80, and the expected heterozygosity from 0.36 to 0.85. Our 10 loci, along with another 12 loci previously described, deliver effective analytical tools to analyse the genetic variability and to assess the social structure of eastern green lizard populations.     

苦荞FtPCS基因克隆、表达及酶活性初步分析

该研究以Pb~(2+)诱导的苦荞(Fagopyrum tataricum)叶片转录组数据为基础,通过RT-PCR克隆,获得苦荞植物络合素合酶(Phytochelatins,PCs)基因(FtPCS);采用无缝克隆构建原核表达载体pET28a-PCS,并通过反向-HPLC结合DTNB[5,5′-二硫代双(2-硝基苯甲酸)]柱后衍生的方法,对纯化的FtPCS重组蛋白在Pb~(2+)存在条件下的催化活性进行初步分析,为进一步揭示FtPCS基因在苦荞重金属富集和解毒过程中的机制奠定基础。结果表明:苦荞FtPCS基因组序列全长5 456bp,包括8个外显子和7个内含子;FtPCS基因ORF序列全长1 485bp,编码494个氨基酸,预测分子量为55.10kDa。可溶性分析表明,苦荞FtPCS基因在E.coli BL21Star(DE3)中以包涵体的形式表达,采用梯度透析复性并结合钴离子螯合层析的方法获得纯化的FtPCS重组蛋白,复性的FtPCS蛋白具有催化GSH生成PC化合物的活性,而且低浓度的Pb~(2+)对其催化活性具有激活作用。     

Development of microsatellite markers and characterization of simple sequence length polymorphism (SSLP) in rice (Oryza sativa L.)

Microsatellite markers containing simple sequence repeats (SSR) are a valuable tool for genetic analysis. Our objective is to augment the existing RFLP map of rice with simple sequence length polymorphisms (SSLP). In this study, we describe 20 new microsatellite markers that have been assigned to positions along the rice chromosomes, characterized for their allelic diversity in cultivated and wild rice, and tested for amplification in distantly related species. Our results indicate that the genomic distribution of microsatellites in rice appears to be random, with no obvious bias for, or clustering in particular regions, that mapping results are identical in intersubspecific and interspecific populations, and that amplification in wild relatives ofOryza sativa is reliable in species most closely related to cultivated rice but becomes less successful as the genetic distance increases. Sequence analysis of SSLP alleles in three relatedindica varieties demonstrated the clustering of complex arrays of SSR motifs in a single 300-bp region with independent variation in each. Two microsatellite markers amplified multiple loci that were mapped onto independent rice chromosomes, suggesting the presence of duplicated regions within the rice genome. The availability of increasing numbers of mapped SSLP markers can be expected to increase the power and resolution of genome analysis in rice.     

Development of microsatellite markers for Quercus miyagii Koidz. (Fagaceae), an endemic species in the Ryukyu Islands, Japan

Quercus miyagii is an endemic tree species in the Ryukyu Islands, Japan. We isolated and characterized 15 microsatellite loci in this species. The number of alleles ranged from 2 to 16 and expected heterozygosities from 0.07 to 0.92. This set of markers is potentially useful to investigate the genetic structure, gene flow, and the biogeographic history of Q. miyagii in the Ryukyu Islands, Japan.     

Isolation and characterization of microsatellite markers in Dysosma versipellis (Berberidaceae), a rare endemic from China

Dysosma versipellis (Berberidaceae) is an important threatened medicinal plant (TMP) species. Here we isolated nine polymorphic microsatellite loci from D. versipellis using a modified biotin-capture method. Our isolated loci provided SSR markers with polymorphism of 2–7 alleles per locus. The expected and observed heterozygosities ranged from 0.507 to 0.864 and from 0.360 to 0.720, respectively. These markers would be the useful tools for analyzing questions concerning population genetic structure and mating system of D. versipellis.     

Development of microsatellite markers for the white mangrove (Laguncularia racemosa C.F. Gaertn., Combretaceae)

We developed nine new microsatellite markers for the white mangrove (Laguncularia racemosa C.F. Gaertn.), a mangrove species widely distributed in the Western Hemisphere. The loci were developed from a sample collected in Baja California Sur, Mexico, and were screened for polymorphism in 108 individuals from the Pacific Coast of Mexico and Costa Rica. The level of polymorphism was relatively low, from two to four alleles. These are the first set of polymorphic markers described for L. racemosa and should be of value in studying regional genetic differentiation among natural populations of this species.     

Development of new microsatellite primers for green and white sturgeon

Sixty-eight primer sets for microsatellite loci were developed from microsatellite motif enriched genomic libraries of pooled DNA from the polyploid green and white sturgeon (Acipenser medirostris and A. transmontanus). Four individuals from each species were screened for polymorphism at these loci. Forty-eight loci amplified in both species, and some exhibited species-specific amplification for white or green sturgeon (8 and 12 loci, respectively). The number of alleles per locus ranged from one to 12. At least 68% of the green and 65% of the white sturgeon loci we developed are polysomic.     

Isolation and characterization of microsatellite loci in Ostryopsis davidiana (Betulaceae)

Ostryopsis (Betulaceae) is a samll genus endemic to China with only two species. Both of them play an important role in restoring the local ecosystems. The distribution of genetic diversity between and within populations in each species are important to further utilize the wild genetic resources and explore the interspecific divergence. In this study, we developed 10 microsatellite loci from O. davidiana by the combining biotin capture method for the first time. A total of 27 microsatellite sequences were recovered through screening the library and 10 of them are polymorphic. The number of alleles per locus in 18 sampled individuals ranged from 3 to 6, expected heterozygosity and observed heterozygosity ranged from 0.2958 to 0.4767 and from 0.1591 to 0.2997, respectively. In addition, all markers have been crossly checked in the other congeneric species. These microsatellite markers would together provide a useful tool for investigating the genetic diversity and structure of both species and speciation mechanism between them.     

Genetic diversity within Oryza rufipogon germplasms preserved in Chinese field gene banks of wild rice as revealed by microsatellite markers

Nineteen microsatellite markers were employed to evaluate the genetic diversity of 92 accessions of common wild rice Oryza rufipogon Griff., which represent a significant portion of the distribution range from field gene banks of China. In comparison, a total of 57 varieties from most of the rice growing areas in China were also analyzed. The microsatellite analysis revealed a considerable amount of genetic diversity resided within the preserved wild rice germplasms. In all, the nineteen microsatellites revealed 328 alleles. The number of alleles per locus varied widely among these markers, ranging from 6 at RM242 to 30 at RM206. A comparison of the genetic parameters showed that wild rice strains preserved in the field gene banks (na = 17.27; R _S = 15.66; H _S = 0.86; H _T = 0.852; H _O = 0.307) possess much higher genetic diversity than cultivated rice varieties (na = 8.27; R _S = 8.14; H _S = 0.75; H _T = 0.758; H _O = 0.051). A total of 196 alleles detected in the wild rice could not be found in cultivated rice, suggesting that about 60% of the alleles of wild rice might be lost during the process of rice domestication. This result shows that these ex situ preserved wild rice strains are of great importance for the discovery and utilization of novel genes in the future rice breeding practices. Considerably abundant genetic variability detected within the studied wild rice germplasms could be comparable to that previously found in a wide sampling of 47 natural populations (na = 16.17; H _S = 0.67; H _O = 0.229), demonstrating that developing field gene banks of wild rice is a necessary and efficient way for preserving genetic diversity of wild rice resources. To determine minimum microsatellites that could distinguish these wild rice accessions, the phylogenetic trees constructed by means of the combinations of different microsatellites suggested that the five highly polymorphic microsatellites could clearly identify these samples. High polymorphisms of rice microsatellite loci and their great resolving power will be particularly helpful for germplasm evaluation and evolutionary studies for better strengthening the conservation and utilization of genetic diversity of wild rice in the field gene banks.     

Isolation and characterization of microsatellite DNA primers in Juniperus przewalskii Kom (Cupressaceae)

Juniperus przewalskii (Cupressaceae) is a dominant tree species endemic to the northeast Qinghai-Tibetan Plateau. This species plays an important role in maintaining the arid ecosystem in this region. However, natural distributions of this species have been declined. In order to develop effective conservation methods, it is important to know the distribution of the genetic diversity within and among populations. In this study, we developed nine new microsatellite loci for this species. We used the combining biotin capture method to enrich AG/CT/AC/GGT microsatellites. The polymorphisms of each locus were further assessed in 12 individuals from four geographically distant populations. The number of alleles per locus varied from three to six and expected heterozygosity ranged from 0.58 to 0.70. These loci together provide a useful tool to investigate the genetic diversity of this species. In addition, all markers have been crossly checked in the other four congeneric species.