Microsatellite marker-mediated analysis of the EMBRAPA Rice Core Collection genetic diversity

The objectives of this study were to determine the genetic structure of 242 accessions from the EMBRAPA Rice Core Collection (ERiCC), to create a mini-core collection and to develop a multiplex panel of fluorescent labeled simple sequence repeats (SSRs). Eighty-six SSRs were used to identify 1,066 alleles, with an average number of 12.4 alleles/locus and average polymorphism information content (PIC)/locus of 0.75. A model-based clustering method recognized the structure of the accessions on two levels, according to their cultivation system and origin. The most divergent subgroup identified was the worldwide lowland accessions, with the highest values for gene diversity (0.75), average Rogers distance modified by Wright (0.80), average number of alleles/locus (11.7) and private alleles (132). A mini-core was assembled with the most divergent 24 lowland and upland accessions. This mini-core displayed an average distance of 0.86, an average number of alleles/locus of 8.4 and an average PIC/locus of 0.8. From the 86 SSRs, 24 were selected to compose six multiplex panels in order to optimize the process of rice genotyping. This set of markers distinguished all 242 accessions, and showed an average PIC of 0.80 and an average number of alleles/locus of 15.4, higher than the entire set of 86 SSRs. Since the heterogeneity found in lines and cultivars of ERiCC was higher than expected, it is necessary to analyze pooled DNA samples to get a better estimate of genetic variability. The SSR characterization of ERiCC clearly indicates that there is high genetic variability in rice accessions stored in genebanks worldwide which can be promptly explored by rice pre-breeding programs.     

Organization of the Rosy Locus in DROSOPHILA MELANOGASTER: Further Evidence in Support of a CIS-Acting Control Element Adjacent to the Xanthine Dehydrogenase Structural Element

The present report summarizes our recent progress in the genetic dissection of an elementary genetic unit in a higher organism, the rosy locus (ry:3--52.0) in Drosophila melanogaster. Pursuing the hypothesis that the rosy locus includes a noncoding control region, as well as a structural element coding for the xanthine dehydrogenase (XDH) peptide, experiments are described that characterize and map a rosy locus variant associated with much lower than normal levels of XDH activity. Experiments are described that fail to relate this phenotype to alteration in the structure of the XDH peptide, but clearly associate this character with variation in number of molecules of XDH per fly. Large-scale fine-structure recombination experiments locate the genetic basis for this variation in the number of molecules of XDH per fly to a site immediately to the left of the XDH structural element within a region previously designated as the XDH control element. Moreover, experiments clearly separate this "underproducer" variant site from a previously described "overproducer" site within the control region. Examination of enzyme activity in electrophoretic gels of appropriate heterozygous genotypes demonstrates the cis-acting nature of this variation in the number of molecules of XDH. A revision of the map of the rosy locus, structural and control elements is presented in the light of the additional mapping data now available.     

Dependence of expected heterozygosity on locus number with stabilizing selection and drift

I determine expected levels of heterozygosity in two allele multilocus models with mutation, stabilizing selection and drift. In the range 2 to 32 loci, the per locus heterozygosity can depend on the locus number. The per locus heterozygosity for ten loci can be as low as three fourths of the per locus heterozygosity in the limit, as the number of loci gets large. Simulations indicate that this dependence on locus number is not due to the population approaching equilibria at which the mean differs from the optimum, but is due to changes in the substitution rate as a function of the number of loci.     

Comparative genomics enabled the isolation of the R3a late blight resistance gene in potato

Comparative genomics provides a tool to utilize the exponentially increasing sequence information from model plants to clone agronomically important genes from less studied crop species. Plant disease resistance (R) loci frequently lack synteny between related species of cereals and crucifers but appear to be positionally well conserved in the Solanaceae. In this report, we adopted a local RGA approach using genomic information from the model Solanaceous plant tomato to isolate R3a, a potato gene that confers race-specific resistance to the late blight pathogen Phytophthora infestans. R3a is a member of the R3 complex locus on chromosome 11. Comparative analyses of the R3 complex locus with the corresponding I2 complex locus in tomato suggest that this is an ancient locus involved in plant innate immunity against oomycete and fungal pathogens. However, the R3 complex locus has evolved after divergence from tomato and the locus has experienced a significant expansion in potato without disruption of the flanking colinearity. This expansion has resulted in an increase in the number of R genes and in functional diversification, which has probably been driven by the co-evolutionary history between P. infestans and its host potato. Constitutive expression was observed for the R3a gene, as well as some of its paralogues whose functions remain unknown.     

普遍野生稻和亚洲栽培稻遗传多样性的研究

用 ４４个 ＲＦＬＰ标记对来自中国、印度、泰国等亚洲 １０个国家的普通野生稻（简称普野,下同）和来自多个国家的７５个栽培稻品种,从多态位点的比率、等位基因数、基因型数、平均杂合度及平均基因多样性等多个方面,比较了不同国家和不同地区的普通野生稻、栽培稻籼粳亚种及栽培稻与普野之间遗传多样性的差异。结果表明：中国普野的遗传多样性最大;其次是印度普野;南亚普野的平均基因多样性大于东南亚普野,而多态位点的比率、等位基因数及基因型数等却低于东南亚普野;栽培稻的遗传多样性明显小于普通野生稻。在所检测的４４个位点中,栽培稻的多态位点数仅为野生稻的３／４,等位基因数约为野生稻的６０％,基因型种类约为野生稻的１／２。栽培稻中籼稻的遗传多样性高于粳稻。在平均每个位点的实际杂合度上,以中国普野杂合度最高,普通野生稻是栽培稻的２倍。说明从野生稻演化成栽培稻的过程中,经过自然选择和人工选择,杂合度降低,等位基因减少,基因多样性下降。     

Linkage disequilibrium mapping of molecular polymorphisms at the scabrous locus associated with naturally occurring variation in bristle number in Drosophila melanogaster

We evaluated the hypothesis that the Drosophila melanogaster second chromosome gene scabrous (sca), a candidate sensory bristle number quantitative trait locus (QTL), contributes to naturally occurring variation in bristle number. Variation in abdominal and sternopleural bristle number was quantified for wild-derived sca alleles in seven genetic backgrounds: as homozygous second chromosomes (C2) in an isogenic background, homozygous lines in which approximately 20 cM including the sca locus had been introgressed into the isogenic background (sca BC), as C2 and sca BC heterozygotes and hemizygotes against a P element insertional sca allele and a P-induced sca deficiency in the same isogenic background, and as sca BC heterozygotes against the wild-type sca allele of isogenic strain. Molecular restriction map variation was determined for a 45 kb region including the sca locus, and single-stranded conformational polymorphism (SSCP) was examined for the third intron and parts of the third and fourth exons. Associations between each of the 27 molecular polymorphisms and bristle number were evaluated within each genotype and on the first principal component score determined from all seven genotypes, separately for each sex and bristle trait. Permutation tests were used to assess the empirical significance thresholds, accounting for multiple, correlated tests, and correlated markers. Three sites in regulatory regions were associated with female-specific variation in abdominal bristle number, one of which was an SSCP site in the region of the gene associated with regulation of sca in embryonic abdominal segments.     

Estimation of copy number in polyploid plants: the good,the bad,and the ugly

Genetic studies in polyploid plants rely heavily on the collection of data from dominant marker loci. A dominant marker locus is a locus for which only the presence or absence of an observable (dominant) allele is recorded. Before these marker loci can be used for genetic exploration, the number of copies of a dominant allele carried by a parent (copy number) must be determined for each marker locus. Copy number in polyploids is estimated using a hypothesis testing procedure. The performance of this estimation procedure has never been evaluated. In this paper, I quantify whether the highly sought after single-copy markers can be accurately identified, if the performance of the estimation procedure improves with increasing sample size, and whether the estimation procedure is capable of accurately estimating the copy number of high copy markers. I found that the probability of incorrectly estimating copy number is quite low and that more data can actually reduce the accuracy of the estimation procedure when the testing assumptions are violated. Fortunately, when a significant result is obtained, it is almost always correct. The challenge often is in obtaining a significant result.     

Linkage mapping of AFLP and microsatellite DNA markers with the body color- and sex-determining loci in the guppy (Poecilia reticulata)

The guppy is an ornamental fish species that exhibits various phenotypic characteristics, such as body color and fin-shape. Although linkage relationships of a limited number of phenotypic traits have already been investigated, the association between phenotypic and molecular markers is still unknown. We constructed a total of 35 linkage groups for the guppy using 186 polymorphic loci of AFLP and microsatellite DNA. The locus related to the yellow body color was linked with ten markers and the sex-determination locus was linked with five markers.     

A small tandem duplication is responsible for the unstable white-ivory mutation in Drosophila

The white-ivory (wi) mutation, an unstable allele of the white locus in Drosophila, reverts to wild-type at frequencies of 5 X 10(-5) in homozygous females, and 5 X 10(-6) in males and deletion heterozygous females. We show by molecular cloning and Southern blot analysis of DNA from wi flies that a 2.9 kilobase tandem duplication within the white locus is responsible for the mutation. Phenotypic reversion appears, in most cases, to be due to an exact excision of the extra copy of the sequence. Two derivative alleles of wi, one phenotypically wild-type, the other a partial revertant, carry insertions of moderately repetitive DNA from outside the locus, in addition to suffering deletions of some white locus DNA. Earlier genetic data preclude unequal crossing-over between homologs as an explanation for the precise reversions. Rather, an intrachromosomal meiotic event seems to be responsible. Our results suggest that intrachromosomal recombination may be responsible in other systems for a larger number of rearrangements than has been suspected, and that interallelic recombination frequencies in Drosophila do not always correlate in a simple way with DNA length or extent of homology.     

A Further Look at Porcine Chromosome 7 Reveals VRTN Variants Associated with Vertebral Number in Chinese and Western Pigs

The number of vertebrae is an economically important trait that affects carcass length and meat production in pigs. A major quantitative trait locus (QTL) for thoracic vertebral number has been repeatedly identified on pig chromosome (SSC) 7. To dissect the genetic basis of the major locus, we herein genotyped a large sample of animals from 3 experimental populations of Chinese and Western origins using 60K DNA chips. Genome-wide association studies consistently identified the locus across the 3 populations and mapped the locus to a 947-Kb region on SSC7. An identical-by-descent sharing assay refined the locus to a 100-Kb segment that harbors only two genes including VRTN and SYNDIG1L. Of them, VRNT has been proposed as a strong candidate of the major locus in Western modern breeds. Further, we resequenced the VRTN gene using DNA samples of 35 parental animals with known QTL genotypes by progeny testing. Concordance tests revealed 4 candidate causal variants as their genotypes showed the perfect segregation with QTL genotypes of the tested animals. An integrative analysis of evolutional constraints and functional elements supported two VRTN variants in a complete linkage disequilibrium phase as the most likely causal mutations. The promising variants significantly affect the number of thoracic vertebrae (one vertebra) in large scale outbred animals, and are segregating at rather high frequencies in Western pigs and at relatively low frequencies in a number of Chinese breeds. Altogether, we show that VRTN variants are significantly associated with the number of thoracic vertebrae in both Chinese and Western pigs. The finding advances our understanding of the genetic architecture of the vertebral number in pigs. Furthermore, our finding is of economical importance as it provides a robust breeding tool for the improvement of vertebral number and meat production in both Chinese indigenous pigs and Western present-day commercial pigs.     

Copy-number fluctuation by unequal crossing-over in the chicken avidin gene family

The chicken avidin gene (AVD) forms a closely clustered gene family together with several avidin-related genes (AVRs). In this study, we used fluorescence in situ hybridization on extended DNA fibers (fiber-FISH) to show that the number of the AVD and AVR genes differs between individuals. Furthermore, the gene copy-number showed wide somatic variation in white blood cells of the individuals. The molecular mechanism underlying the fluctuation is most probably unequal crossing-over and/or unequal sister chromatid exchange, as judged by the Gaussian distribution of the gene counts. By definition, an increase in gene number on one locus should be accompanied by a decrease on the other locus in unequal sequence exchange. The results suggest that copy-number lability may be more common among gene families than previously thought. The chicken avidin gene family also provides an excellent model for studying the mechanisms of recombination and gene conversion.     

The probability distribution under a population divergence model of the number of genetic founding lineages of a population or species

The composition of genetic variation in a population or species is shaped by the number of events that led to the founding of the group. We consider a neutral coalescent model of two populations, where a derived population is founded as an offshoot of an ancestral population. For a given locus, using both recursive and nonrecursive approaches, we compute the probability distribution of the number of genetic founding lineages that have given rise to the derived population. This number of genetic founding lineages is defined as the number of ancestral individuals that contributed at the locus to the present-day derived population, and is formulated in terms of interspecific coalescence events. The effects of sample size and divergence time on the probability distribution of the number of founding lineages are studied in detail. For 99.99% of the loci in the derived population to each have one founding lineage, the two populations must be separated for 9.9N generations. However, only approximately 0.87N generations must pass since divergence for 99.99% of the loci to have <6 founding lineages. Our results are useful as a prior expectation on the number of founding lineages in scenarios that involve the evolution of one population from the splitting of an ancestral group, such as in the colonization of islands, the formation of polyploid species, and the domestication of crops and livestock from wild ancestors.     

Intragenic suppression: Stalker,a retrovirus-like transposable element,can compensate for a deficiency at the cut locus of Drosophila melanogaster

A number of mutations at the cut locus were induced by non-precise exision of a silent P-element insertion which resulted in deletions at the regulatory region of the locus. Unexpectedly, a reversion of one of these mutations was found, which appears as a result of insertion of Stalker (a retrovirus-like mobile element) near the 1.3 kb deletion. Thus an insertion of a retrovirus-like mobile element can suppress the deficiency at the regulatory region of a gene.     

A multipoint linkage map of the distal short arm of the human X chromosome.

The distal portion of the short arm of the human X chromosome (Xp) exhibits many unique and interesting features. Distal Xp contains the pseudoautosomal region, a number of disease loci, and several cell-surface markers. Several genes in this area have also been observed to escape X-chromosomal inactivation. The characterization of new polymorphic loci in this region has permitted the construction of a refined multipoint linkage map extending 15 cM from the Xp telomere. This interval is known to contain the loci for the diseases X-linked ichthyosis, chondrodysplasia punctata, and Kallmann syndrome, as well as the cell-surface markers Xg and 12E7. This region also contains the junction between the pseudoautosomal region and strictly X-linked sequences. The locus MIC2 has been demonstrated by linkage analysis to be indistinguishable from the pseudoautosomal junction. The steroid sulfatase locus has been mapped to an interval adjacent to the DXS278 locus and 6 cM from the pseudoautosomal junction. The polymorphic locus (STS) DXS278 was shown to be informative in all families studied, and linkage analysis reveals that the locus represents a low-copy repeat with at least one copy distal to the STS gene. The generation of a multipoint linkage map of distal Xp will be useful in the genetic dissection of many of the unique features of this region.     

Real-time PCR for the detection of precise transgene copy number in durum wheat

Recent results obtained in various crops indicate that real-time PCR could be a powerful tool for the detection and characterization of transgene locus structures. The determination of transgenic locus number through real-time PCR overcomes the problems linked to phenotypic segregation analysis (i.e. lack of detectable expression even when the transgenes are present) and can analyse hundreds of samples in a day, making it an efficient method for estimating gene copy number. Despite these advantages, many authors speak of “estimating” copy number by real-time PCR, and this is because the detection of a precise number of transgene depends on how well real-time PCR performs.     

The use marker alleles for the introgression of linked quantitative alleles

Summary It is shown that when an exotic strain and a commercial strain differ genetically at a quantitative locus and at an adjoining marker locus, repeated backcrosses to the commercial strain, retaining only backcross progeny carrying the exotic marker allele, will allow the effective introgression of the linked quantitative allele from the exotic to the commercial strain. The introgression procedure will be particularly effective when exotic and commercial strains differ at two nearby marker loci with the quantitative locus bracketed between them. The simultaneous introgression of a number of quantitative alleles from different exotic strains, and appropriate selection procedures in the intercross generations that follow are also considered.     

Joining the loops: beta-globin gene regulation

The mammalian beta-globin locus is a multigene locus containing several globin genes and a number of regulatory elements. During development, the expression of the genes changes in a process called "switching." The most important regulatory element in the locus is the locus control region (LCR) upstream of the globin genes that is essential for high-level expression of these genes. The discovery of the LCR initially raised the question how this element could exert its effect on the downstream globin genes. The question was solved by the finding that the LCR and activate globin genes are in physical contact, forming a chromatin structure named the active chromatin hub (ACH). Here we discuss the significance of ACH formation, provide an overview of the proteins implicated in chromatin looping at the beta-globin locus, and evaluate the relationship between nuclear organization and beta-globin gene expression.     

Unequivocal determination of sire allele origin for multiallelic microsatellites when only the sire and progeny are genotyped

The effect of a segregating economic trait locus (ETL) can be detected with the aid of a linked genetic marker, if specific alleles of each locus are in association among the individuals genotyped for the genetic marker. For dairy cattle this can be achieved by application of the ‘granddaughter design’. If only the sires and their sons are genotyped for the genetic markers, then the allele origin of sons having the same genotypes as their sires cannot be determined. Seven sires and 101 sons were genotyped for five microsatellites. The mean frequency of heterozygous sires was 77%. The mean number of alleles per locus was 8.2. Frequency of informative sons per locus ranged from 60% to 80% with a mean of 72%. With highly polymorphic microsatellites, at least 60% more grandsire families can be included in the analysis, and the number of sons assayed can be reduced by 40%, as compared to diallelic markers.     

Structure of the catfish IGH locus: analysis of the region including the single functional IGHM gene

The catfish IGH locus is large (∼1 Mb) and complex, having undergone multiple internal duplications and transpositions. To define the structure of the locus that contains the single expressed IGHM gene, two overlapping bacterial-artificial-chromosome (BAC) clones spanning the most 3′ end of the channel catfish immunoglobulin heavy (IGH) chain locus have been completely sequenced. The analyses created a contig of 257,153 bp containing 55 VH, 6 D, 12 JH genes and the IGH constant region genes encoding the functional secreted and membrane forms of IgM and the membrane form of IgD. This analysis revealed three major features. First, no C-region genes were found aside from the previously described IGHM1 and IGHD1, with the latter gene being the most 3′ C-region gene of the catfish IGH locus. There was no evidence in the region sequenced for genes that could encode an Ig class similar to the IgZ/IgT described in zebrafish, trout and pufferfish. Second, there are a high number of VH pseudogenes, 28 out of 55 (51%). In contrast, the entire zebrafish IGH locus has 40 functional VH genes and eight pseudogenes (17%). Third, an internal duplication of a 52.4-kb block of VH genes has occurred. These observations suggest that the IGH locus of teleost fish varies significantly from species to species in the diversity of C-region genes as well as the numbers of genes encoding V regions.