Cloning of AFLP markers linked to resistance to Peronosclerospora sorghi in maize

Genetic mapping of resistance genes for sorghum downy mildew (SDM) in maize revealed multiple-locus inheritance. A combination of AFLP (amplified fragment length polymorphism) technique with bulked segregant analysis (BSA) was applied to map the genes involved in the resistance to SDM (Peronosclerospora sorghi) in a recombinant inbred population. Three AFLP markers were identified and mapped to chromosomes 1 and 9, in regions previously associated with SDM resistance. One other AFLP marker was found to be associated with disease susceptibility but could not be linked to any chromosome. These four AFLP fragments were isolated, cloned and sequenced. A BLAST search of the GenBank database showed that none of these four sequences was closely related to resistance genes that have been reported previously. Sequence-characterized amplified regions (SCARs) were produced and used to assess the presence of SDM resistance genes and characterize specific genotypes. These markers may be useful in marker-assisted breeding programs.     

Splenic gene expression profiling in White Leghorn layer inoculated with the Salmonella enterica serovar Enteritidis

Salmonella enterica serovar Enteritidis (SE) is a foodborne pathogen that can threaten human health through contaminated poultry products. Live poultry, chicken eggs and meat are primary sources of human salmonellosis. To understand the genetic resistance of egg‐type chickens in response to SE inoculation, global gene expression in the spleen of 20‐week‐old White Leghorn was measured using the Agilent 4 × 44 K chicken microarray at 7 and 14 days following SE inoculation (dpi). Results showed that there were 1363 genes significantly differentially expressed between inoculated and non‐inoculated groups at 7 dpi (I7/N7), of which 682 were up‐regulated and 681 were down‐regulated genes. By contrast, 688 differentially expressed genes were observed at 14 dpi (I14/N14), of which 371 were up‐regulated genes and 317 were down‐regulated genes. There were 33 and 28 immune‐related genes significantly differentially expressed in the comparisons of I7/N7 and I14/N14 respectively. Functional annotation revealed that several Gene Ontology (GO) terms related to immunity were significantly enriched between the inoculated and non‐inoculated groups at 14 dpi but not at 7 dpi, despite a similar number of immune‐related genes identified between I7/N7 and I14/N14. The immune response to SE inoculation changes with different time points following SE inoculation. The complicated interaction between the immune system and metabolism contributes to the immune responses to SE inoculation of egg‐type chickens at 14 dpi at the onset of lay. GC, TNFSF8, CD86, CD274, BLB1 and BLB2 play important roles in response to SE inoculation. The results from this study will deepen the current understanding of the genetic response of the egg‐type chicken to SE inoculation at the onset of egg laying.     

Identification of Loci Associated with Drought Resistance Traits in Heterozygous Autotetraploid Alfalfa (Medicago sativa L.) Using Genome-Wide Association Studies with Genotyping by Sequencing

Drought resistance is an important breeding target for enhancing alfalfa productivity in arid and semi-arid regions. Identification of genes involved in drought tolerance will facilitate breeding for improving drought resistance and water use efficiency in alfalfa. Our objective was to use a diversity panel of alfalfa accessions comprised of 198 cultivars and landraces to identify genes involved in drought tolerance. The panel was selected from the USDA-ARS National Plant Germplasm System alfalfa collection and genotyped using genotyping by sequencing. A greenhouse procedure was used for phenotyping two important traits associated with drought tolerance: drought resistance index (DRI) and relative leaf water content (RWC). Marker-trait association identified nineteen and fifteen loci associated with DRI and RWC, respectively. Alignments of target sequences flanking to the resistance loci against the reference genome of M. truncatula revealed multiple chromosomal locations. Markers associated with DRI are located on all chromosomes while markers associated with RWC are located on chromosomes 1, 2, 3, 4, 5, 6 and 7. Co-localizations of significant markers between DRI and RWC were found on chromosomes 3, 5 and 7. Most loci associated with DRI in this work overlap with the reported QTLs associated with biomass under drought in alfalfa. Additional significant markers were targeted to several contigs with unknown chromosomal locations. BLAST search using their flanking sequences revealed homology to several annotated genes with functions in stress tolerance. With further validation, these markers may be used for marker-assisted breeding new alfalfa varieties with drought resistance and enhanced water use efficiency.     

Conservation of the syntenies between porcine chromosome 7 and human chromosomes 6, 14 and 15 demonstrated by radiation hybrid mapping and linkage analysis

Comparative mapping studies facilitate the identification of genes located in quantitative trait locus (QTL) regions in domestic animals by utilizing information from the human genome. Radiation hybrid (RH) mapping is effective for this purpose because of its high resolution in ordered gene mapping on chromosomes. We constructed an RH map of pig chromosome 7, by adding 23 markers associated with genes. This RH map clearly demonstrated the mosaic of homology between pig chromosome 7 (SSC7) and human chromosomes 6, 14 and 15 at a 'gene' level, and was confirmed by linkage analysis. Clarification of the homology of SSC7 to human chromosomes will contribute to the elucidation of the gene(s) responsible for QTL detected on this chromosome.     

Comparative mapping of human chromosome 10 to pig chromosomes 10 and 14

Identification of predictive markers in QTL regions that impact production traits in commercial populations of swine is dependent on construction of dense comparative maps with human and mouse genomes. Chromosomal painting in swine suggests that large genomic blocks are conserved between pig and human, while mapping of individual genes reveals that gene order can be quite divergent. High-resolution comparative maps in regions affecting traits of interest are necessary for selection of positional candidate genes to evaluate nucleotide variation causing phenotypic differences. The objective of this study was to construct an ordered comparative map of human chromosome 10 and pig chromosomes 10 and 14. As a large portion of both pig chromosomes are represented by HSA10, genes at regularly spaced intervals along this chromosome were targeted for placement in the porcine genome. A total of 29 genes from human chromosome 10 were mapped to porcine chromosomes 10 (SSC10) and 14 (SSC14) averaging about 5 Mb distance of human DNA per marker. Eighteen genes were assigned by linkage in the MARC mapping population, five genes were physically assigned with the IMpRH mapping panel and seven genes were assigned on both maps. Seventeen genes from human 10p mapped to SSC10, and 12 genes from human 10q mapped to SSC14. Comparative maps of mammalian species indicate that chromosomal segments are conserved across several species and represent syntenic blocks with distinct breakpoints. Development of comparative maps containing several species should reveal conserved syntenic blocks that will allow us to better define QTL regions in livestock.     

Genetics of preeclampsia: an approach to genetic linkage studies

Due to its high prevalence during pregnancies, preeclampsia is considered an important public health problem. Many investigators agree in that its expression is related to the interaction between genetic and environmental factors. Many studies have searched for genetic factors, attempting to identify chromosomal regions or candidate genes whose variants may be related to high preeclampsia susceptibility. Several studies have associated a number of susceptibility genes to preeclampsia, but the results have not been replicated consistently in all populations. Mapping of genes and chromosomal regions by linkage analysis has located potential markers on chromosomes 2 and 4. Identification of the genes located in these candidate regions will pinpoint the genetic risk factors, will lead to a better understanding of the syndrome, and will provide clues for its prevention and treatment.     

Porcine genomics delivers new tools and results: this little piggy did more than just go to market

The past decade has yielded new tools for pig geneticists and breeders thanks to the considerable developments resulting from efforts to map the pig genome. The pig genetic linkage map now has nearly 5000 loci including several hundred genes, microsatellites and amplified fragment length polymorphisms (AFLP) markers. Using tools that include somatic cell hybrid panels and radiation hybrid panels, the physical genetic map is also growing rapidly and has over 4000 genes and markers. Scientists using both exotic and commercial breeds for quantitative trait loci (QTL) scans and candidate gene analyses have identified a number of important chromosomal regions and individual genes associated with growth rate, leanness, feed intake, meat quality, litter size and disease resistance. Using marker-assisted selection (MAS) the commercial pig industry is actively incorporating these gene markers and traditional performance information to improve traits of economic importance in pig production. Researchers now have novel tools including pig gene arrays and advanced bioinformatics that are being exploited to find new candidate genes and to advance the understanding of gene function in the pig. Sequencing of the pig genome has been initiated and further sequencing is now being considered. Advances in pig genomics and directions for future research and the implications to both the pig industry and human health are reviewed.     

Identification of genes controlling collagen-induced arthritis in mice: striking homology with susceptibility loci previously identified in the rat.

The susceptibility to collagen-induced arthritis in the highly susceptible DBA/1 mouse has earlier been shown to be partly controlled by the MHC class II gene Aq. To identify susceptibility loci outside of MHC, we have made crosses between DBA/1 and the less susceptible B10.Q strain, both expressing the MHC class II gene Aq. Analysis of 224 F2 intercross mice with 170 microsatellite markers in a genome-wide scan suggested 4 quantitative trait loci controlling arthritis susceptibility located on chromosomes 6, 7, 8, and 10. The locus on chromosome 6 (Cia6), which was associated with arthritis onset, yielded a logarithm of odds score of 4.7 in the F2 intercross experiment and was reproduced in serial backcross experiments. Surprisingly, the DBA/1 allele had a recessive effect leading to a delay in arthritis onset. The suggestive loci on chromosomes 7 and 10 were associated with arthritis severity rather than onset, and another suggestive locus on chromosome 8 was most closely associated with arthritis incidence. The loci on chromosomes 7, 8, and 10 all appeared to contain disease-promoting alleles derived from the DBA/1 strain. Interestingly, most of the identified loci were situated in chromosomal regions that are homologous to regions in the rat genome containing susceptibility genes for arthritis; the mouse Cia6 locus is homologous with the rat Cia3, Pia5, Pia2, and Aia3; the locus on chromosome 7 (Cia7) is homologous with the rat Cia2; and the locus on chromosome 10 (Cia8) is homologous with the rat Cia4.     

Deletion-based physical mapping of barley chromosome 7H

Chromosomal mutations in barley (Hordeum vulgare, 2n=2x=14, HH) chromosome 7H added to the common wheat (Triticum aestivum, 2n=6x=42, AABBDD) cultivar Chinese Spring were induced genetically by the gametocidal activity of certain alien chromosomes derived from wild species of the genus Aegilops. The rearranged barley chromosomes were characterized by C-banding, FISH and GISH. Twenty two deletion or translocation chromosomes in a hemizygous condition were selected for deletion mapping of 17 AFLP and 28 STS markers that are specific to 7H. Of the 22 breakpoints in chromosome 7H, seven involved the short arm (7HS), 12 the long arm (7HL) and three were in the centromeric region. The seven 7HS breakpoints separated all four 7HS-specific AFLP markers and split the 21 STS markers into six groups. One breakpoint occurred between two STS markers formerly occupying the same position in the genetic map. All seven 7HS breakpoints were separated from each other by either the AFLP or STS markers. The 12 breakpoints in 7HL divided the 13 7HL-specific AFLP markers into seven groups, and the seven STS markers into three groups. On the other hand, the 12 breakpoints in 7HL were divided into six groups by the AFLP markers and into two groups by the STS markers. This deletion-based map was in accordance with previously published genetic and physical maps using the same STS markers. The breakpoints, AFLP markers and STS markers were arrayed in a consistent order. Received: 5 February 2001 / Accepted: 19 February 2001     

Advances in pig genomics and functional gene discovery

Advances in pig gene identification, mapping and functional analysis have continued to make rapid progress. The porcine genetic linkage map now has nearly 3000 loci, including several hundred genes, and is likely to expand considerably in the next few years, with many more genes and amplified fragment length polymorphism (AFLP) markers being added to the map. The physical genetic map is also growing rapidly and has over 3000 genes and markers. Several recent quantitative trait loci (QTL) scans and candidate gene analyses have identified important chromosomal regions and individual genes associated with traits of economic interest. The commercial pig industry is actively using this information and traditional performance information to improve pig production by marker-assisted selection (MAS). Research to study the co-expression of thousands of genes is now advancing and methods to combine these approaches to aid in gene discovery are under way. The pig's role in xenotransplantation and biomedical research makes the study of its genome important for the study of human disease. This review will briefly describe advances made, directions for future research and the implications for both the pig industry and human health.     

From a sow's ear to a silk purse: real progress in porcine genomics

An incredible amount of progress has occurred in the past decade since the pig genome map began to develop. The porcine genetic linkage map now has nearly 5,000 loci including several hundred genes, microsatellites and amplified fragment length polymorphism (AFLP) markers being added to the map. Thanks to somatic cell hybrid panels and then radiation hybrid panels the physical genetic map is also growing rapidly and now has over 4,000 genes and markers. Many quantitative trait loci (QTL) scans have been completed and together with candidate gene analyses have identified important chromosomal regions and individual genes associated with traits of economic interests. Using marker assisted selection (MAS) the commercial pig industry is actively using this information and traditional performance information to improve pig production. Large scale pig arrays are just now beginning to be used and co-expression of thousands of genes is now advancing our understanding of gene function. The pig's role in xenotransplantation and biomedical research makes the study of its genome important for the study of human disease. Sequencing of the pig genome appears on the near horizon. This commentary will discuss recent advances in pig genomics, directions for future research and the implications to both the pig industry and human health.     

Mapping of sex determination loci on the white campion (Silene latifolia) Y chromosome using amplified fragment length polymorphism

S. latifolia is a dioecious plant with morphologically distinct sex chromosomes. To genetically map the sex determination loci on the male-specific Y chromosome, we identified X-ray-induced sex determination mutants that had lost male traits. We used male-specific AFLP markers to characterize the extent of deletions in the Y chromosomes of the mutants. We then compared overlapping deletions to predict the order of the AFLP markers and to locate the mutated sex-determining genes. We found three regions on the Y chromosome where frequent deletions were significantly associated with loss of male traits. One was associated with hermaphroditic mutants. A second was associated with asexual mutants that lack genes needed for early stamen development and a third was associated with asexual mutants that lack genes for late stages of stamen development. Our observations confirmed a classical genetic prediction that S. latifolia has three dispersed male-determining loci on the Y chromosome, one for carpel suppression, one for early stamen development, and another for late stamen development. This AFLP map provides a framework for locating genes on the Y chromosome and for characterizing deletions on the Y chromosomes of potentially interesting mutants.     

A genetic linkage map for coho salmon (Oncorhynchus kisutch)

Construction of genetic linkage maps is an important first step for a variety of genomic applications, such as selective breeding in aquaculture, comparative studies of chromosomal evolution and identification of loci that have played key roles in the evolution of a species. Here we present a sex-specific linkage map for coho salmon. The map was constructed using 148 AFLP markers, 133 microsatellite loci and the phenotypic locus SEX . Twenty-four linkage groups spanning 287.4 cM were mapped in males, and 33 linkage groups spanning 429.7 cM were mapped in females. Several male linkage groups corresponded to two female linkage groups. The combination of linkage groups across both sexes appeared to characterize regions of 26 chromosomes. Two homeologous chromosomes were identified based on information from duplicated loci. Homologies between the coho and rainbow trout maps were examined. Eighty-six loci were found to form common linkage relationships between the two maps; these relationships provided evidence for whole-arm fissions, fusions and conservation of chromosomal regions in the evolution of these two species.     

Beyond an AFLP genome scan towards the identification of immune genes involved in plague resistance in Rattus rattus from Madagascar

Genome scans using amplified fragment length polymorphism (AFLP) markers became popular in nonmodel species within the last 10 years, but few studies have tried to characterize the anonymous outliers identified. This study follows on from an AFLP genome scan in the black rat (Rattus rattus), the reservoir of plague (Yersinia pestis infection) in Madagascar. We successfully sequenced 17 of the 22 markers previously shown to be potentially affected by plague‐mediated selection and associated with a plague resistance phenotype. Searching these sequences in the genome of the closely related species Rattus norvegicus assigned them to 14 genomic regions, revealing a random distribution of outliers in the genome (no clustering). We compared these results with those of an in silico AFLP study of the R. norvegicus genome, which showed that outlier sequences could not have been inferred by this method in R. rattus (only four of the 15 sequences were predicted). However, in silico analysis allowed the prediction of AFLP markers distribution and the estimation of homoplasy rates, confirming its potential utility for designing AFLP studies in nonmodel species. The 14 genomic regions surrounding AFLP outliers (less than 300 kb from the marker) contained 75 genes encoding proteins of known function, including nine involved in immune function and pathogen defence. We identified the two interleukin 1 genes (Il1a and Il1b) that share homology with an antigen of Y. pestis, as the best candidates for genes subject to plague‐mediated natural selection. At least six other genes known to be involved in proinflammatory pathways may also be affected by plague‐mediated selection.     

Genome-wide scan for bovine twinning rate QTL using linkage disequilibrium

Twinning is a complex trait with negative impacts on health and reproduction, which cause economic loss in dairy production. Several twinning rate quantitative trait loci (QTL) have been detected in previous studies, but confidence intervals for QTL location are broad and many QTL are unreplicated. To identify genomic regions or genes associated with twinning rate, QTL analysis based on linkage combined with linkage disequilibrium (LLD) and individual marker associations was conducted across the genome using high-throughput single nucleotide polymorphism (SNP) genotypes. A total of 9919 SNP markers were genotyped with 200 sires and sons in 19 half-sib North American Holstein dairy cattle families. After SNPs were genotyped, informative markers were selected for genome-wide association tests and QTL searches. Evidence for twinning rate QTL was found throughout the genome. Thirteen markers significantly associated with twinning rate were detected on chromosomes 2, 5 and 14 ( P  < 2.3 × 10⁻⁵). Twenty-six regions on fourteen chromosomes were identified by LLD analysis at P  < 0.0007. Seven previously reported ovulation or twinning rate QTL were supported by results of single marker association or LLD analyses. Single marker association analysis and LLD mapping were complementary tools for the identification of putative QTL in this genome scan.     

Assignment of 64 genes expressed in 28-day-old pig embryo to radiation hybrid map

A swine resource family was constructed at the National Institute of Animal Industry, Japan, in order to determine the genetic regions responsible for economically important traits, including fetus development. To identify genes expressed in the early stage of embryo development, we cataloged and mapped genes expressed in a 28-day-old normal pig embryo. In this effort, we have mapped 64 genes, which have map information in human genome onto a swine radiation hybrid (RH) map, IMpRH. These mappings provided additional chromosomal homologies between swine and human to improve the comparative map between the two species. The distribution of the genes assigned to swine chromosomes are as follows: 9 genes were assigned on SSC6; 6 genes each assigned on SSC5 and SSC14; 5 genes each assigned on SSC3, SSC4, and SSC8; 4 genes each assigned on SSC1, SSC7, SSC9, and SSC15; 3 genes each assigned on SSC2, SSC13 and SSCX; and 1 gene each assigned on SSC10, SSC11, and SSC16. Moreover, the present findings revealed 18 new chromosomal homologies between pig and human. Briefly, SSC3 regions were indicated to correspond with HSA1 and HSA10; SSC4 with HSA6; SSC5 with HSA2, HSA15, and HSA16; SSC6 with HSA3, HSA6, and HSA20; SSC7 with HSA11; SSC8 with HSA3, HSA6, and HSA7; SSC9 with HSA8; SSC13 with HSA1; SSC14 with HSA13; SSC15 with HSA19; SSC16 with HSA9. Received: 19 December 2000 / Accepted: 13 March 2001     

An AFLP-based genetic linkage map of Plasmodium chabaudi chabaudi

Background

Plasmodium chabaudi chabaudi can be considered as a rodent model of human malaria parasites in the genetic analysis of important characters such as drug resistance and immunity. Despite the availability of some genome sequence data, an extensive genetic linkage map is needed for mapping the genes involved in certain traits.

Methods

The inheritance of 672 Amplified Fragment Length Polymorphism (AFLP) markers from two parental clones (AS and AJ) of P. c. chabaudi was determined in 28 independent recombinant progeny clones. These, AFLP markers and 42 previously mapped Restriction Fragment Length Polymorphism (RFLP) markers (used as chromosomal anchors) were organized into linkage groups using Map Manager software.

Results

614 AFLP markers formed linkage groups assigned to 10 of 14 chromosomes, and 12 other linkage groups not assigned to known chromosomes. The genetic length of the genome was estimated to be about 1676 centiMorgans (cM). The mean map unit size was estimated to be 13.7 kb/cM. This was slightly less then previous estimates for the human malaria parasite, Plasmodium falciparum

Conclusion

The P. c. chabaudi genetic linkage map presented here is the most extensive and highly resolved so far available for this species. It can be used in conjunction with the genome databases of P. c chabaudi, P. falciparum and Plasmodium yoelii to identify genes underlying important phenotypes such as drug resistance and strain-specific immunity.     

Chromosomal localization of homeobox genes and associated markers on porcine Chromosomes 3, 5, 12, 15, 16 and 18: comparative mapping study with human and mouse

Four homeobox genes that belong to the four homeobox gene clusters known in mammals have been regionally assigned to four distinct porcine chromosomes in conserved regions between human and pig. HOXA11, HOXB6, HOXC8, and HOXD4 genes were mapped by radioactive in situ hybridization to porcine Chromosomes (Chrs) 18q21-24 (with a secondary signal in 16q14-21), 12p11-12, 5p11-12, and 15q22-23 respectively. Besides, we have also revealed the presence of a porcine homeobox (pig Hbx24) which, although showing DNA sequence homology with a mouse gene of HOXB cluster, was located on porcine Chr 3 (3p14-13) outside the Hox clusters. To support the identity of the homeobox gene clusters analyzed and in the light of the high sequence similarity among homeobox genes, we also localized markers known to be mapped near each Hox cluster in human. In this way, four genes were also mapped in pig: GAPD (5q12-21), GAD1 (15q21-22), INHBA (18q24), and IGFBP3 (18q24). Mapping of HOXA11, INHBA, and IGFBP3 on pig Chr 18 constitutes the first assignments of genes on this small chromosome. These new localizations extend the information on the conservation of four human chromosomal regions in the pig genome. Received: 7 August 1995 / Accepted: 16 October 1995