Evaluation of the genetic profile of the Pulawska breed

An assessment was made of the genetic variation of the Pulawska pig through the determination of polymorphism of 6 genes and 14 microsatellite sequences. The examinations covered 52 gilts included in a preservation breeding project. The identification of the alleles at microsatellite loci was performed in an ABI PRISM 310 GENETIC ANALYZER. Gene polymorphism was established by the PCR-RLFP method. On the basis of the variation of 6 genes and 14 microsatellites the mean value of the heterozygosity coefficient was estimated at 0.61, while the value of the corresponding PIC coefficient (polymorphism information content) amounted to 0.55. The probability that the genotypes of two randomly chosen individuals in a population are identical was: 6.95 x 10(-3) (based on gene allele frequency) and 1.23 x 10(-14) (based on microsatellite allele frequency).     

Multiplexed systems of microsatellite markers for genetic analysis of mahogany,Swietenia macrophylla King (Meliaceae), a threatened neotropical timber species

Mahogany (Swietenia macrophylla King [Meliaceae]) is the most valuable hardwood species in the neotropics. Its conservation status has been the subject of increasing concern due to overexploitation and habitat destruction. In this work we report the development and characterization of 10 highly variable microsatellite loci for S. macrophylla. Twenty-nine percent of the 126 sequenced mahogany clones yielded useful microsatellite loci. Three high-throughput genotyping systems were developed based on polymerase chain reaction (PCR) multiplexing of these mahogany loci. We identified a total of 158 alleles in 121 adult individuals of S. macrophylla, with an average of 15.8 alleles (range 11-25) per locus. All loci showed Mendelian inheritance in open-pollinated half-sib families. The mean expected heterozygosity was 0.84 and the mean observed heterozygosity was 0.73. The combined probability of identity-the probability that two individuals selected at random from a population would have identical genotypes--was 7.0 x 10(-15), and combined probability of paternity exclusion was 0.999998 overall loci. These microsatellite loci permit precise estimates of parameters such as gene flow, mating system, and paternity, thus providing important insights into the population genetics and conservation of S. macrophylla.     

Analysis of 22 heterologous microsatellite markers for genetic variability in Indian goats

The genetic variability of 22 heterologous microsatellite markers was analyzed in two Indian goat breeds, namely Bengal and Chegu. The heterozygosity, polymorphism information content (PIC), and probability of identity of two individuals were calculated for all microsatellite loci in both the breeds. The observed number of alleles varied between 4 and 13 at the studied microsatellite loci. The evaluated microsatellite loci exhibited high mean heterozygosity of 0.69 +/- 0.11 and 0.66 +/- 0.07 in Bengal and Chegu goats, respectively. The mean PIC values of the studied loci in these breeds were 0.79 +/- 0.08 and 0.78 +/- 0.05, respectively. The probability of identity of two random individuals from different breeds, taking into account, all the 22 microsatellite loci was as low as 5.523 x 10(-40). On the basis of these results, we propose that these microsatellite markers may be used with reliability for studying genetic diversity and for identification of individuals in Indian goat breeds.     

Assessment of genetic variation and differentiation of hop genotypes by microsatellite and AFLP markers.

Microsatellites have many desirable marker properties and have been increasingly used in crop plants in genetic diversity studies. Here we report on the characterisation of microsatellite markers and on their use for the determination of genetic identities and the assessment of genetic variability among accessions from a germplasm collection of hop. Thirty-two polymorphic alleles were found in the 55 diploid genotypes, with an average number of eight alleles (3.4 effective alleles) for four microsatellite loci. Calculated polymorphic information content values classified three loci as informative markers and two loci as suitable for mapping. The average observed heterozygosity was 0.7 and the common probability of identical genotypes was 3.271 x 10(-4). An additional locus, amplified by one primer pair, was confirmed by segregation analysis of two crosses. The locus discovered was heterozygous, with a null allele in the segregating population. The same range of alleles was detected in nine triploid and five tetraploid hop genotypes. Cultivar heterozygosity varied among all 69 accessions, with only one cultivar being homozygous at four loci. Microsatellite allele polymorphisms distinguished 81% of all genotypes; the same allelic profile was found mainly in clonally selected cultivars. Cultivar-specific alleles were found in some genotypes, as well as a specific distribution of alleles in geographically distinct hop germplasms. The genetic relationship among 41 hop accessions was compared on the basis of microsatellite and AFLP polymorphisms. Genetic similarity dendrograms showed low correlation between the two marker systems. The microsatellite dendrogram grouped genetically related accessions reasonably well, while the AFLP dendrogram showed good clustering of closely related accessions and, additionally, separated two geographically distinct hop germplasms. The results of microsatellite and AFLP analysis are discussed from the point of view of the applicability of the two marker systems for different aspects of germplasm evaluation.     

雌核发育银鲫微卫星突变速率和突变模式

银鲫是天然雌核发育的三倍体两性型种群, 因其遗传背景和生殖方式的特殊性, 已经成为研究单性和多倍体脊椎动物进化遗传学的理想模式鱼类。利用33个微卫星序列对雌核发育银鲫的突变速率和突变模式进行研究, 结果表明: (1) 22个子代个体中, 检测到1尾个体在15个微卫星位点具有18个突变等位基因; (2) 每个微卫星位点每代总体平均突变率是1.16×10-2, 95%置信区间是6.87×10-3~1.83×10-2, 与其他鱼类相比, 雌核发育银鲫的突变率明显偏高, 这与天然雌核发育鱼类处在单性生殖和两性生殖的过度阶段有密切关系; (3) 具有突变等位基因的13个位点的重复单元数目都在10次以上, 11个复合型微卫星位点的突变率(1.31×10-2)与21个完美型位点(1.00×10-2)的突变率没有明显差异(P = 0.67), 微卫星突变率受到重复单元数目的影响, 然而与重复结构类型和侧翼序列GC碱基含量无相关性; (4) 序列分析表明, 雌核发育银鲫的微卫星突变模式并不严格遵守逐步突变模型。     

Microsatellite Loci Polymorphism of Apple (<Emphasis Type="Italic">Malus domestica</Emphasis> Borkh.) Genotypes with Different Ploidy Level

In this paper, the microsatellite (SSR) loci analysis was used to study apple genotypes with different levels of ploidy. A total of 47 samples were studied (9 diploids, 21 triploids, and 17 tetraploids) for seven microsatellite loci (GD147, Hi02C07, CH02c11, CH04c07, CH03d07, CH02c09, and GD12). It was possible to refine the pedigrees for some forms. It was established that the tetraploidss 20-9-30 and 20-9-27, selected in a hybrid family from the crossing of Wealthy 4x and Antonovka Obyknovennaya, were probably obtained from the self-pollination of the maternal form, since in the most loci they did not inherit alleles from the paternal form. As a result of the alleles distribution analysis, the spontaneous triploid cultivars Nizkorosloe and Sinap Orlovsky were revealed to be formed from the merge of an unreduced ovum and haploid pollen, since in the heterozygous loci both alleles are inherited from the maternal form and only one from the paternal form. According to the obtained data, studied tetraploids may be divided into two groups, which also reflect the features of tetraploids origin. The first group includes tetraploids inherited alleles from one initial diploid form (including spontaneous and induced tetraploids, as well as forms from self-pollination of the tetraploid maternal form). These teraploids, like diploids, amplify 1–2 alleles per locus (on average, for all 7 loci, one genotype amplifies 13 alleles). The second group includes tetraploids carrying alleles from several initial diploid forms. Tetraploids of this group are highly heterozygous and amplify 3–4 alleles at most loci (the maximum number of alleles at all loci, 24 alleles, was identified in the form 30-47-88). Tetraploids of the second group have a greater potential for the genetic diversity of its offspring. Analysis of polymorphism of microsatellite loci can be used (1) as an alternative or additional method for identifying the triploid hybrids from heteroploid crosses of orthoploid forms, which is based on the analysis of the loci most polymorphic in parental forms, and (2) for the analysis of true hybridity (verification of pedigrees), including tetraploid forms. Moreover, we identified the most polymorphic loci suitable for the above purposes. The aspects of qualitative and quantitative interpretation of the fragment analysis of microsatellite loci results are considered. The possibilities and limitations of the SSR analysis for detection of apple ploidy level are discussed.     

Polymorphic microsatellite loci for the mustached tamarin (Saguinus mystax) and their cross‐species amplification in other New World monkeys

We isolated and characterized 10 polymorphic microsatellite loci for the mustached and cotton‐top tamarin (Saguinus mystax and S. oedipus) and also tested their applicability in other New World monkey genera. Six loci proved to be variable in Cebus apella and Saimiri sciureus/S. boliviensis, respectively. We also show that faeces‐derived DNA is suitable for polymerase chain reaction (PCR) analyses of a primate taxon that gives birth to twins with individuals exhibiting a haematopoetic chimaerism. The microsatellite fingerprint patterns do not display a complex mixture of the PCR‐products derived from the alleles of both dizygotic twins, but exhibit the constitutive alleles of one sibling only.     

Characteristics of loci and individuals are associated with germline microsatellite mutation rates in lesser kestrels (Falco naumanni)

Although microsatellites are one of the most popular tools in genetic studies, their mutational dynamics and evolution remain unclear. Here, we apply extensive pedigree genotyping to identify and analyze the patterns and factors associated with de novo germline mutations across nine microsatellite loci in a wild population of lesser kestrels (Falco naumanni). A total of 10 germline mutations events were unambiguously identified in four loci, yielding an average mutation rate of 2.96x10(-3). Across loci, mutation rate was positively correlated with locus variability and average allele size. Mutations were primarily compatible with a stepwise mutation model, although not exclusively involved single-step changes. Unexpectedly, we found an excess of maternally transmitted mutations (male-to-female ratio of 0.1). One of the analyzed loci (Fn2.14) resulted hypermutable (mutation rate=0.87%). This locus showed a size-dependent mutation bias, with longer alleles displaying deletions or additions of a small number of repeat than shorter alleles. Mutation probability at Fn2.14 was higher for females and increased with parental (maternal) age but was not associated with individual physical condition, multilocus heterozygosity, allele length or allele span. Overall, our results do not support the male-biased mutation rate described in other organisms and suggest that mutation dynamics at microsatellite loci are a complex process which requires further research.     

凡纳滨对虾微卫星位点在两个选育家系中遗传的初步研究

利用两个选育凡纳滨对虾全同胞家系研究了10个微卫星位点的遗传特征。通过ABI310或3100测序仪检测, 在所观察到的20个基因型比例（genotypic ratios）(10个微卫星位点 X 2个家系)中,有17个基因型比例符合孟德尔遗传。微卫星位点TUMXLv8.220在两个家系中均存在无效等位基因,从而3个不符合孟德尔遗传基因型中2个可由无效等位基因来解释。TUMXLv 3.1在06家系偏离了1：1：1：1的孟德尔预期比。3个微卫星位点（TUMXLv5.66,TUMXLv7.74,TUMXLv8.224）在两个家系中均表现单态。3个微卫星位点（TUMXLv5.45,TUMXLv7.56,TUMXLv8.256）在两个家系均既表现多态又遵循孟德尔共显性遗传, 是亲子鉴定和种群遗传分析的较好选择。结果显示在应用微卫星标记进行遗传分析之前利用全同胞家系进行遗传模式研究是非常必要的。     

Multiple paternity and female-biased mutation at a microsatellite locus in the olive ridley sea turtle (Lepidochelys olivacea)

Multiple paternity in the olive ridley sea turtle (Lepidochelys olivacea) population nesting in Suriname was demonstrated using two microsatellite loci, viz., Ei8 and Cm84. The large number of offspring sampled per clutch (70 on average, ranging from 15 to 103) and the number of alleles found at the two loci (18 and eight alleles, respectively) enabled unambiguous assessment of the occurrence of multiple paternity. In two out of 10 clutches analysed, the offspring had been sired by at least two males, which was confirmed at both loci. In both clutches, unequal paternity occurred: 73% and 92% of the offspring had been sired by the primary male. The probability of detecting multiple paternity was 0.903, and therefore there is a small chance that multiple paternity occurred but remained undetected in some of the eight clutches that appeared to be singly sired. Analysis of 703 offspring revealed a high mutation rate for locus Ei8 (micro = 2.3 x 10(-2)) with all 33 mutations occurring in maternal alleles. In particular, one allele of 274 bp mutated at a high frequency in a clutch to which the mother contributed the allele, but in another clutch where the father contributed the same allele, no such mutations were observed. Inferred allele-specific mutation rates for Ei8 and expected numbers of mutations per clutch confirmed that maternal alleles for Ei8 are more likely to mutate in the olive ridley sea turtle than paternal alleles. Possible explanations are discussed.     

High mutation rate of a long microsatellite allele in Drosophila melanogaster provides evidence for allele-specific mutation rates

Within recent years, microsatellite have become one of the most powerful genetic markers in biology. For several mammalian species, microsatellite mutation rates have been estimated on the order of 10(- 3)-10(-5). A recent study, however, demonstrated mutation rates in Drosophila melanogaster of at least one order of magnitude lower than those in mammals. To further test this result, we examined mutation rates of different microsatellite loci using a larger sample size. We screened 24 microsatellite loci in 119 D. melanogaster lines maintained for approximately 250 generations and detected 9 microsatellite mutations. The average mutation rate of 6.3 x 10(-6) is identical to the mutation rate from a previous study. Most interestingly, all nine mutations occurred at the same allele of one locus (DROYANETSB). This hypermutable allele has 28 dinucleotide repeats and is among the longest microsatellite reported in D. melanogaster. The allele-specific mutation rate of 3.0 x 10(-4) per generation is within the range of mammalian mutation rates. Future microsatellite analyses will have to account for the dramatic differences in allele-specific mutation rates.      

秦川母牛群体遗传特性的微卫星标记研究

为了从DNA分子水平揭示秦川牛群体遗传多态性和群体遗传结构,寻找可用于秦川牛的微卫星标记,本研究选择了12个普通牛（Bos taurus）微卫星标记检测了90头秦川母牛各微卫星位点的遗传变异及多态性。结果表明,在秦川母牛群体中,12个微卫星位点共检测到了247个等位基因,各位点的等位基因数在13（INRA005）～33个（HEL13）之间,平均每个微卫星位点的等位基因数为21个;总有效等位基因数和平均每个位点平均有效等位基因数（Ne）分别分为142．6229和11．8852。各位点平均基因频率取样方差（V（pij））为2．6036×10^-4。12个微卫星位点平均观察杂合度（Ho）和平均期望杂合度（He）在0．7842（INRA005）～0．9775（BM315）和0．7952（BM315）～0．9446（HEL13）之间。12个位点平均多态信息含量（PIC）在0．7653（INRA005）～0．9420（HEL13）之间,平均为0．8965．12个微卫星位点均属于高度多态位点,这表明秦川母牛群体中所检测各微卫星位点具有丰富的遗传多态性,具备较大的选择潜力。12个微卫星位点的平均固定指数（F）为-0．0076,即各位点杂合子的缺陷度不高,即偏离Hardy—Weinberg平衡的程度不大。     

Mutation in microsatellite repeats of DNA and embryonal death in humans

In the analysis of tetranucleotide DNA repeats inheritance carried out in 55 families with a history of spontaneous miscarriages and normal karyotypes in respect to 21 loci located on seven autosomes, 8 embryos (14.5%) demonstrating 12 cases of the presence of alleles absent in both parents were described. The study of chromosome segregation using other DNA markers permitted highly probable exclusion of false paternity as well as uniparental disomy as the reasons for parent/child allele mismatches. The high probability of paternity together with the presence of a "new" allele at any offspring locus points to the mutation having occurred during game-togenesis in one of the parents. Examination of mutation in spontaneous abortuses revealed an increased number of tandem repeat units at microsatellite loci in three cases and an decreased number of these repeats in six cases. In two abortuses, a third allele absent in both parents, which resulted from a somatic mutation that occurred during embryonic development, was observed. The prevalence of the male germline mutations, revealed during investigation of the mutation origin, was probably associated with an increased number of DNA replication cycles in sperm compared to the oocytes. In spontaneous abortuses, the mean mutation rate of the tetranucleotide repeat complexes analyzed was 9.8 x 10(-3) per locus per gamete per generation. This was about five times higher than the spontaneous mutation rate of these STR loci. It can be suggested that genome instability detected at the level of repeated DNA sequences can involve not only genetically neutral loci but also active genomic regions crucial for embryonic viability. This results in cell death and termination of embryonic development. Our findings indicate that the death of embryos with normal karyotypes in most cases is associated with an increased frequency of germline and somatic microsatellite mutations. The data of the present study also provide a practical tool for the quantitative evaluation of this phenomenon and for the analysis of the reasons for miscarriages and embryonic death in certain families.     

Development and characterization of 15 novel microsatellite markers from forest musk deer (<Emphasis Type="Italic">Moschus berezovskii</Emphasis>)

A total of 15 polymorphic microsatellite loci were isolated from forest musk deer (Moschus berezovskii). These loci showed high levels of allelic diversity, presenting 6–14 alleles per locus in a group with 31 individuals. The mean observed and expected heterozygosities were 0.82 and 0.68, respectively. The average polymorphic information content value, cumulative discrimination power, overall probability of exclusion with both unknown parents or with only unknown sire for these markers were 0.78, 1.0, 0.9999 and 1.0, respectively, indicating that this suit of microsatellite loci would be a powerful tool for the genetic determination of the forest musk deer.     

Genetic diversity and relationships among Dutch elm disease tolerant Ulmus pumila L. accessions from China.

Elm breeding programs worldwide have relied heavily on Asian elm germplasm, particularly Ulmus pumila, for the breeding of Dutch elm disease tolerant cultivars. However, the extent and patterning of genetic variation in Asian elm species is unknown. Therefore, the objective of this research was to determine the extent of genetic diversity among 53 U. pumila accessions collected throughout the People's Republic of China. Using 23 microsatellite loci recently developed in the genus Ulmus, a total of 94 alleles were identified in 15 polymorphic and 4 monomorphic loci. The average number of alleles per locus was 4.9, with a range of 1-11 alleles. Gene diversity estimates per locus ranged from 0.08 to 0.87, and the non-exclusion probability for the 15 polymorphic loci combined was 0.7 x 10(-9). Nineteen region-specific alleles were identified, and regional gene diversity estimates were moderately high (0.48-0.57). The genetic relationships among accessions and regions were estimated by UPGMA and principal coordinate analysis. Both techniques discriminated all accessions and regions. Two microsatellite markers (UR175 + UR123 or Ulm-3) were sufficient to discriminate up to 99.7% of the accessions studied. This research provides useful information for DNA-based fingerprinting, breeding, ecological studies, and diversity assessment of elm germplasm.     

Polymorphism of bovine microsatellite DNA sequences in the lowland European bison

Investigations of genetic polymorphism of microsatellite DNA sequences were conducted in 22 individuals of the European bisonBison bonasus (Linneaus, 1758) from Bia?owie?a Primeval Forest. For this purpose 27 cattle microsatellite primer pairs were used. Among the 27 microsatellite markers examined, an amplification product was obtained for 21 loci. This rendered it possible to identify total of 40 alleles in the bison population tested. In addition, eight loci were proved to be monomorphic. A majority of the 40 alleles identified was identical with the alleles identified at the corresponding loci in cattle. Only two alleles seem to be specific for the European bison. The value of heterozygosity for the examined loci in bison population from Bia?owie?a was low and ranged from 0.13 to 0.53. Hence, the polymorphism information content was low as well. Based on our results the microsatellite DNA markers identified in cattle may be used to analyse the genetic structure of the population of European bison.     

The effective mutation rate at Y chromosome short tandem repeats, with application to human population-divergence time

We estimate an effective mutation rate at an average Y chromosome short-tandem repeat locus as 6.9x10-4 per 25 years, with a standard deviation across loci of 5.7x10-4, using data on microsatellite variation within Y chromosome haplogroups defined by unique-event polymorphisms in populations with documented short-term histories, as well as comparative data on worldwide populations at both the Y chromosome and various autosomal loci. This value is used to estimate the times of the African Bantu expansion, the divergence of Polynesian populations (the Maoris, Cook Islanders, and Samoans), and the origin of Gypsy populations from Bulgaria.     

An Evaluation of Evolutionary Constraints on Microsatellite Loci Using Null Alleles

A test to evaluate constraints on the evolution of single microsatellite loci is described. The test assumes that microsatellite alleles that share the same flanking sequence constitute a series of alleles with a common descent that is distinct from alleles with a mutation in the flanking sequence. Thus two or more different series of alleles at a given locus represent the outcomes of different evolutionary processes. The higher rate of mutations within the repeat region (10(-3) or 10(-4)) compared with that of insertion/deletion or point mutations in adjacent flanking regions (10(-9)) or with that of recombination between the repeat and the point mutation (10(-6) for sequences 100 bp long) provides the rationale for this assumption. Using a two-phase, stepwise mutation model we simulated the evolution of a number of independent series of alleles and constructed the distributions of two similarity indices between pairs of these allele series. Applying this approach to empirical data from locus AG2H46 of Anopheles gambiae resulted in a significant excess of similarity between the main and the null series, indicating that constraints affect allele distribution in this locus. Practical considerations of the test are discussed.     

Microsatellite DNA marker inheritance indicates preferential pairing between two highly homologous genomes in polyploid and hemisexual dog-roses, Rosa L. Sect. Caninae DC

According to previous cytological evidence, the hemisexual dog-rose species, Rosa sect. Caninae, transmit only seven chromosomes (derived from seven bivalents) through their pollen grains, whereas egg cells contain 21, 28 or 35 chromosomes (derived from seven bivalents and 14, 21 or 28 univalents) depending on ploidy level. Two sets of reciprocal pairwise interspecific crosses involving the pentaploid species pair R. dumalis and R. rubiginosa, and the pentaploid/tetraploid species pair R. sherardii and R. villosa, were analysed for 13 and 12 microsatellite DNA loci, respectively. Single loci were represented by a maximum of three simultaneously occurring alleles in R. villosa, and four alleles in the other three parental plants. In the experimentally derived offspring, the theoretical maximum of five alleles was found for only one locus in the pentaploid progenies. Microsatellite DNA allele composition was identical with that of the maternal parent in 10 offspring plants, which were probably derived through apomixis. Almost all microsatellite DNA alleles were shared with the maternal parent also in the remaining offspring, but 1-4 alleles shared only with the paternal parent, indicating sexual seed formation. Analysis of quantitative peak differences allowed a tentative estimation of allelic configuration in the individual plants, and suggested that bivalent formation preferentially takes place between chromosomes that consistently share the same microsatellite alleles and therefore appear to be highly homologous. Moreover, alleles that were shared between the species in each cross combination comparatively often appear to reside on the bivalent-forming chromosomes, whereas species-specific alleles instead occur comparatively often on the univalent-forming chromosomes and are therefore inherited through the maternal parent only. Recombination then takes place between very similar genomes also in interspecific crosses, resulting in a reproduction system that is essentially a mixture between apomixis and selfing.     

Statistical genetic considerations for maintaining germ plasm collections

One objective of the regeneration of genetic populations is to maintain at least one copy of each allele present in the original population. Genetic diversity within populations depends on the number and frequency of alleles across all loci. The objectives of this study on outbreeding crops are: (1) to use probability models to determine optimal sample sizes for the regeneration for a number of alleles at independent loci; and (2) to examine theoretical considerations in choosing core subsets of a collection. If we assume that k-1 alleles occur at an identical low frequency of p₀ and that the k^th allele occurs at a frequency of 1-[(k-1)p₀], for loci with two, three, or four alleles, each with a p₀ of 0.05, 89–110 additional individuals are required if at least one allele at each of 10 loci is to be retained with a 90% probability; if 100 loci are involved, 134–155 individuals are required. For two, three, or four alleles, when p₀ is 0.03 at each of 10 loci, the sample size required to include at least one of the alleles from each class in each locus is 150–186 individuals; if 100 loci are involved, 75 additional individuals are required. Sample sizes of 160–210 plants are required to capture alleles at frequencies of 0.05 or higher in each of 150 loci, with a 90–95% probability. For rare alleles widespread throughout the collection, most alleles with frequencies of 0.03 and 0.05 per locus will be included in a core subset of 25–100 accessions.