Mapping of 443 porcine EST improves the comparative maps for SSC1 and SSC7 with the human genome

Numerous mapping studies of complex traits in the pig have resulted in quantitative trait loci (QTL) intervals of 10-20 cM. To improve the chances to identify the genes located in such intervals, increased expressed sequence tags (EST)-based marker density, coupled with comparative mapping with species whose genomes have been sequenced such as human and mouse, is the most efficient tool. In this study, we mapped 443 porcine EST with a radiation hybrid (RH) panel (384 had LOD > 6.0) and a somatic cell hybrid panel. Requiring no discrepancy between two-point and multipoint RH data allowed robust assignment of 309 EST, of which most were located on porcine chromosomes (SSC) 1, 4, 7, 8 and X. Moreover, we built framework maps for two chromosomes, SSC1 and SSC7, with mapped QTL in regions with known rearrangement between pig and human genomes. Using the Blast tool, we found orthologies between 407 of the 443 pig cDNA sequences and human genes, or to existing pig genes. Our porcine/human comparative mapping results reveal possible new homologies for SSC1, SSC3, SSC5, SSC6, SSC12 and SSC14 and add markers in synteny breakpoints for chromosome 7.     

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.     

绵羊基因组研究进展

过去几年中,家畜基因组计划取得了巨大进展。已经构建了猪、鸡、牛、绵羊、马、鹿的遗传图谱,其遗传标记间距在5～20cM。这些图谱对于家畜中与重要经济性状相关的基因或遗传标记的鉴定非常重要。该文从绵羊的基因图谱、比较图谱、重要经济性状基因及QTL定位方面对绵羊基因组的研究进展作了简要阐述。 Abstract:During the last few years,advances in livestock genome projects have been remarkable.Species-specific genetic maps exist for pig,chicken,cattle,sheep,horse,and deer with marker intervals of 5 to 20 cM.These maps have been essential for the identification of genes and genetic markers associated with importantly economic traits in livestock.In this paper,advances of gene map,comparative map,the genes for importantly economic traits and quantitative trait loci (QTL) mapping were briefly introduced in sheep.     

Comparative mapping of a region on chromosome 10 containing QTL for reproduction in swine

Several quantitative trait loci (QTL) for important reproductive traits (age of puberty, ovulation rate, nipple number and plasma FSH) have been identified on the long arm of porcine chromosome 10. Bi-directional chromosome painting has shown that this region is homologous to human chromosome 10p. Because few microsatellite or type I markers have been placed on SSC10, we wanted to increase the density of known ESTs mapped in this region of the porcine genome. Genes were chosen for their position on human chromosome 10, sequence availability from the TIGR pig gene indices, and their potential as a candidate gene. The PCR primers were designed to amplify across introns or 3'-UTR to maximize single nucleotide polymorphism (SNP) discovery. Parents of the mapping population (one sire and seven dams) were amplified and sequenced to find informative markers. The SNPs were genotyped using primer extension and mass spectrometry. These amplification products were also used to probe a BAC library (RPCI-44, Roswell Park Cancer Institute) for positive clones and screened for microsatellites. Six genes from human chromosome 10p (AKR1C2, PRKCQ, ITIH2, ATP5C1, PIP5K2A and GAD2) were mapped in the MARC swine mapping population. Gene order was conserved within these markers from centromere to telomere of porcine chromosome 10q, as compared with human chromosome 10p. Four of these genes (PIP5K2A, ITIH2, GAD2 and AKR1C2), which map under QTL, are potential candidate genes. Identification of porcine homologues near important QTL and development of a comparative map for this chromosome will allow further fine- mapping and positional cloning of candidate genes affecting reproductive traits.     

Linkage mapping of gene-associated SNPs to pig chromosome 11

Single nucleotide polymorphisms (SNPs) were discovered in porcine expressed sequence tags (ESTs) orthologous to genes from human chromosome 13 (HSA13) and predicted to be located on pig chromosome 11 (SSC11). The SNPs were identified as sequence variants in clusters of EST sequences from pig cDNA libraries constructed in the Sino-Danish pig genome project. In total, 312 human gene sequences from HSA13 were used for similarity searches in our pig EST database. Pig ESTs showing significant similarity with HSA13 genes were clustered and candidate SNPs were identified. Allele frequencies for 26 SNPs were estimated in a group of 80 unrelated pigs from Danish commercial pig breeds: Duroc, Hampshire, Landrace and Large White. Eighteen of the 26 SNPs genotyped in the PiGMaP Reference Families were mapped by linkage analysis to SSC11. The EST-based SNPs published here are new genetic markers useful for linkage and association studies in commercial and experimental pig populations. This study represents the first gene-associated SNP linkage map of pig chromosome 11 and adds new comparative mapping information between SSC11 and HSA13. Furthermore, our data facilitate future studies aimed at the identification of interesting regions on pig chromosome 11, positional cloning and fine mapping of quantitative trait loci in pig.     

A successful strategy for comparative mapping with human ESTs: 65 new regional assignments in the pig

Large-scale sequencing of cDNAs from numerous tissues is currently being performed within the framework of the Human Genome Project. These expressed sequence tags (ESTs) are then mapped on a radiation hybrid panel to produce a high-resolution map of human genes. In this report, we estimate the efficiency of mapping these ESTs in the pig. A total of 344 human ESTs from Généthon were selected for amplification in other species by Zoo-PCR: 186 of these could be reproducibly amplified by use of pig DNA and the corresponding human primer pairs. One-hundred seven of these were tested on a porcine–rodent somatic cell hybrid panel, permitting regional localizations of 65 ESTs with agarose or single-strand conformation polymorphism analysis gels. The corresponding pig PCR products were sequenced: 60 ESTs matched significantly with the expected human sequences. Fifty-one of these localizations in the pig are in agreement with the comparative mapping data between humans and pigs based on heterologous chromosome painting. Seven ESTs that were localized in an unexpected region may indicate new chromosomal correspondences. This work significantly increases the number of genes mapped on the pig genome and demonstrates that this approach can be successfully applied to improve the gene density of mammalian genomic maps in chromosomal regions of interest, such as those in which QTL (Quantative Trait Loci) have been identified. Received: 31 July 1998 / Accepted: 14 October 1998     

Construction of a dense SNP map for bovine chromosome 6 to assist the assembly of the bovine genome sequence

A linkage map was constructed for bovine chromosome 6 (BTA6), using 399 single nucleotide polymorphisms (SNPs) detected primarily from PCR-resequencing. The efficiency of SNP detection was highly dependent on the source of sequence information chosen for primer design (BAC-end sequences, introns or promoters). The SNPs were used to build a linkage map comprising 104 cM on BTA6. The SNP order in the linkage map corresponded very well with radiation hybrid (RH) maps available for BTA6 as well as with expected positions in the human comparative map, but diverged significantly from the current assembly of the bovine genome (Btau_3.1). When performing linkage analysis with the marker order suggested from the Btau_3.1 we observed an expansion of the genetic map from 104 cM to 137 cM, strongly suggesting a reordering of scaffolds in the current version of the bovine genome assembly. The extent of LD on BTA6 was evaluated by calculating the average r ² for SNP pairs separated by given distances. The decline of LD was rapid with distance, such that r ² was 0.1 at 100 kb. Our results indicate that linkage mapping will be a valuable source of information for correcting errors in the current bovine assembly. These errors were sufficiently frequent to be of concern for the accuracy of mapping QTL with panels of SNPs whose positions are based on the current assembly.     

Improving the comparative map of SSC2p-q13 by sample sequencing of BAC clones

To improve the comparative map for pig chromosome 2 and increase the gene density on this chromosome, a porcine bacterial artificial chromosome (BAC) library was screened with 17 microsatellite markers and 18 genes previously assigned to pig chromosome 2. Fifty-one BAC clones located in the region of a maternally imprinted quantitative trait locus for backfat thickness (BFT) were identified. From these BACs 372 kb were sample sequenced. The average read length of a subclone was 442 basepair (bp). Contig assembly analysis showed that every bp was sequenced 1.28 times. Subsequently, sequences were compared with sequences in the nucleotide databases to identify homology with other mammalian sequences. Sequence identity was observed with sequences derived from 35 BACs. The average percentage identity with human sequences was 87.6%, with an average length of 143 bp. In total, sample sequencing of all BACs resulted in sequence identity with 29 human genes, 13 human expressed sequence tags (ESTs), 17 human genomic clones, one rat gene, one porcine gene and nine porcine ESTs. Eighteen genes located on human chromosome 11 and 19, and seven genes from other human locations, one rat gene and one porcine gene were assigned to pig chromosome 2 for the first time. The new genes were added to the radiation hybrid map at the same position as the locus from which the BAC that was sequenced was derived. In total 57 genes were placed on the radiation hybrid map of SSC2p-q13.     

Investigation of Obesity Candidate Genes On Porcine Fat Deposition Quantitative Trait Loci Regions

Objectives: To investigate possible obesity candidate genes in regions of porcine quantitative trait loci (QTL) for fat deposition and obesity‐related phenotypes. Research Methods and Procedures: Chromosome mapping and QTL analyses of obesity candidate genes were performed using DNA panels from a reference pig family. Statistical association analyses of these genes were performed for fat deposition phenotypes in several other commercial pig populations. Results: Eight candidate genes were mapped to QTL regions of pig chromosomes in this study. These candidate genes also served as anchor loci to determine homologous human chromosomal locations of pig fat deposition QTL. Preliminary analyses of relationships among polymorphisms of individual candidate genes and a variety of phenotypic measurements in a large number of pigs were performed. On the basis of available data, gene‐gene interactions were also studied. Discussion: Comparative analysis of obesity‐related genes in the pig is not only important for development of marker‐assisted selection on growth and fat deposition traits in the pig but also provides for an understanding of their genetic roles in the development of human obesity.     

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.     

ETph: enhancers and their targets in pig and human database

Enhancers, as the genomic non-coding sequences, play a key role in the activation of gene expression. They have been widely identified in the human genome. Pig is an important biomedical model for human health. Few studies have been performed to explore the enhancers in the pig genome. The human enhancer information may be useful to identify enhancers in the pig genome. In addition, the genetic background of pig traits could be useful to annotate human enhancers and diseases. Thus, in order to further study enhancers and their potential roles in human and pig, we developed a public database, ETph (Enhancers and their Targets in pig and human). ETph integrates the information on human enhancers, pig putative enhancers, target genes, pig QTL terms, human diseases, GO terms and the KEGG pathway. A total of 25 182 enhancers were identified in the pig genome using the human homology sequence information. Among them, 6232 high-confidence enhancers were used to build the ETph. ETph provides a convenient platform to search, browse and download data. Moreover, a web-based analytical tool was designed to visualize networks and topology graphs among pig putative enhancers, target genes, pig QTL traits and human diseases. ETph might provide a useful tool for researchers to investigate the genetic background of pig traits and human diseases. ETph is freely accessible at http://klab.sjtu.edu.cn/enhancer/ .     

Copy number variation detection using SNP genotyping arrays in three Chinese pig breeds

We performed genome‐wide CNV detection based on SNP genotyping data of 96 Chinese‐native Tibetan, Dahe and Wuzhishan pigs. These pigs are particularly interesting because of their excellent adaptation to hypoxia or small body size, which facilitates the use of them as models of different human diseases in addition to valuable agricultural animals. A total of 105 CNV regions (CNVRs) were identified, encompassing 16.71 Mb of the pig genome. Seven of 10 (70%) CNVRs selected randomly were validated by quantitative real‐time PCR. Comparison with previous studies revealed 25 (23.81%) novel CNVRs, indicating that CNV coverage of the pig genome is still incomplete and there exists large diversity between pig breeds. Functional analysis of genes located in these CNVRs confirmed the high representation of genes involved in sensory perception, neurological system processes and other basic metabolic processes. In addition, the majority of these CNVRs were detected to span reported pig QTL that affect various traits, which highlighted three biologically interesting genes with copy number changes (i.e., ANKRD34B, FAM110B and ABCG1). These genes may have economic importance in pig breeding and are worth being further investigated. We also obtained some CNVRs harboring genes that had human orthologs involved in human diseases such as cardiovascular disease and Alzheimer's disease. The findings of this study are a significant extension of the coverage of CNVRs in the pig genome and provide valuable resources for follow‐up‐associated studies of CNVs in pig complex traits as well as important implications of human diseases.     

A potato molecular-function map for carbohydrate metabolism and transport

Molecular-linkage maps based on functional gene markers (molecular-function maps) are the prerequisite for a candidate-gene approach to identify genes responsible for quantitative traits at the molecular level. Genetic linkage between a quantitative trait locus (QTL) and a candidate-gene locus is observed when there is a causal relationship between alleles of the candidate gene and the QTL effect. Functional gene markers can also be used for marker-assisted selection and as anchors for structural and functional comparisons between distantly related plant species sharing the same metabolic pathways. A first molecular-function map with 85 loci was constructed in potato based on 69 genes. Priority was given to genes operating in carbohydrate metabolism and transport. Public databases were searched for genes of interest from potato, tomato, or other plant species. DNA sequence information was used to develop PCR-based marker assays that allowed the localization of corresponding potato genes on existing RFLP linkage maps. Comparing the molecular-function map for genes operating in carbohydrate metabolism and transport with a QTL map for tuber starch content indicates a number of putative candidate genes for this important agronomic trait. Received: 19 March 2000 / Accepted: 16 May 2000     

Gains in QTL detection using an ultra-high density SNP map based on population sequencing relative to traditional RFLP/SSR markers

Huge efforts have been invested in the last two decades to dissect the genetic bases of complex traits including yields of many crop plants, through quantitative trait locus (QTL) analyses. However, almost all the studies were based on linkage maps constructed using low-throughput molecular markers, e.g. restriction fragment length polymorphisms (RFLPs) and simple sequence repeats (SSRs), thus are mostly of low density and not able to provide precise and complete information about the numbers and locations of the genes or QTLs controlling the traits. In this study, we constructed an ultra-high density genetic map based on high quality single nucleotide polymorphisms (SNPs) from low-coverage sequences of a recombinant inbred line (RIL) population of rice, generated using new sequencing technology. The quality of the map was assessed by validating the positions of several cloned genes including GS3 and GW5/qSW5, two major QTLs for grain length and grain width respectively, and OsC1, a qualitative trait locus for pigmentation. In all the cases the loci could be precisely resolved to the bins where the genes are located, indicating high quality and accuracy of the map. The SNP map was used to perform QTL analysis for yield and three yield-component traits, number of tillers per plant, number of grains per panicle and grain weight, using data from field trials conducted over years, in comparison to QTL mapping based on RFLPs/SSRs. The SNP map detected more QTLs especially for grain weight, with precise map locations, demonstrating advantages in detecting power and resolution relative to the RFLP/SSR map. Thus this study provided an example for ultra-high density map construction using sequencing technology. Moreover, the results obtained are helpful for understanding the genetic bases of the yield traits and for fine mapping and cloning of QTLs.     

小麦旗叶早衰性状的QTL定位

为小麦旗叶早衰性状的精细定位和基因克隆奠定基础,该试验以普通小麦(Triticum aestivum L.)‘宁春4号’和‘宁春27号’杂交得到的128个F10代RIL群体为研究材料,利用307对多态性SSR标记对小麦旗叶早衰性状进行了QTL定位,并通过构建整合图谱的方法进行了标记加密。结果表明,共检测到1个控制旗叶早衰性状的加性QTL,位于2A染色体长臂的gwm526和gwm382标记区间内,可解释49.88%的表型变异。经遗传图谱整合后发现,gwm526和gwm382标记之间存在124个SNP标记。     

Investigation of a QTL region for loin eye area and fatness on pig Chromosome 1

Previously, quantitative trait loci (QTL) for tenth-rib backfat (TENTHRIB) and loin eye area (LEA) were identified on pig Chromosome 1 (SSC 1) near microsatellite S0008 from a three-generation Berkshire × Yorkshire cross (BY). This work attempted to refine these QTL positions and identify genes associated with these QTL. Genotypes of BY (n = 555) were determined by PCR-RFLP or PCR tests for 13 polymorphisms identified in BY F₀ individuals for candidate genes, BAC end sequences, and genomic clones. Using least-squares regression interval mapping, the LEA QTL was estimated at S0008; the TENTHRIB QTL position was shifted approximately 1 cM downstream from S0008. Of the genes/sequences mapped in the QTL region, CL349415 was significantly associated with TENTHRIB (p = 0.02) and solute carrier family 2, member 12 (SLC2A12) was significantly associated with LEA (p = 0.02). These results suggest that the gene(s) responsible for the LEA and TENTHRIB QTL effects are tightly linked to S0008 or that the high informativeness of S0008 relative to surrounding markers is influencing the QTL position estimates. In addition, janus kinase 2 (JAK2) was mapped to a suggestive LEA QTL region and showed association with LEA (p = 0.009), fatness, color, and pH traits in BY.     

First-generation SNP/InDel markers tagging loci for pathogen resistance in the potato genome

A panel of 17 tetraploid and 11 diploid potato genotypes was screened by comparative sequence analysis of polymerase chain reaction (PCR) products for single nucleotide polymorphisms (SNPs) and insertion-deletion polymorphisms (InDels), in regions of the potato genome where genes for qualitative and/or quantitative resistance to different pathogens have been localized. Most SNP and InDel markers were derived from bacterial artificial chromosome (BAC) insertions that contain sequences similar to the family of plant genes for pathogen resistance having nucleotide-binding-site and leucine-rich-repeat domains (NBS-LRR-type genes). Forty-four such NBS-LRR-type genes containing BAC-insertions were mapped to 14 loci, which tag most known resistance quantitative trait loci (QTL) in potato. Resistance QTL not linked to known resistance-gene-like (RGL) sequences were tagged with other markers. In total, 78 genomic DNA fragments with an overall length of 31 kb were comparatively sequenced in the panel of 28 genotypes. 1498 SNPs and 127 InDels were identified, which corresponded, on average, to one SNP every 21 base pairs and one InDel every 243 base pairs. The nucleotide diversity of the tetraploid genotypes (pi = 0.72 x 10(-3)) was lower when compared with diploid genotypes (pi = 2.31 x 10(-3)). RGL sequences showed higher nucleotide diversity when compared with other sequences, suggesting evolution by divergent selection. Information on sequences, sequence similarities, SNPs and InDels is provided in a database that can be queried via the Internet.     

Construction of a high-density,high-resolution genetic map and its integration with BAC-based physical map in channel catfish

Construction of genetic linkage map is essential for genetic and genomic studies. Recent advances in sequencing and genotyping technologies made it possible to generate high-density and high-resolution genetic linkage maps, especially for the organisms lacking extensive genomic resources. In the present work, we constructed a high-density and high-resolution genetic map for channel catfish with three large resource families genotyped using the catfish 250K single-nucleotide polymorphism (SNP) array. A total of 54,342 SNPs were placed on the linkage map, which to our knowledge had the highest marker density among aquaculture species. The estimated genetic size was 3,505.4 cM with a resolution of 0.22 cM for sex-averaged genetic map. The sex-specific linkage maps spanned a total of 4,495.1 cM in females and 2,593.7 cM in males, presenting a ratio of 1.7 : 1 between female and male in recombination fraction. After integration with the previously established physical map, over 87% of physical map contigs were anchored to the linkage groups that covered a physical length of 867 Mb, accounting for ∼90% of the catfish genome. The integrated map provides a valuable tool for validating and improving the catfish whole-genome assembly and facilitates fine-scale QTL mapping and positional cloning of genes responsible for economically important traits.     

Animal QTLdb: beyond a repository

Over the past ten years there have been a large number of publications that have described hundreds of quantitative trait loci (QTL) in livestock species. To facilitate the comparison of QTL results across experiments, the Animal QTL database (QTLdb) was developed to house all published QTL information as a public repository. The QTLdb was originally developed to serve the porcine genomics community (previously known as PigQTLdb). We have further developed the QTLdb to house QTL data from multiple species, including but not limited to cattle, chickens, and pigs. In addition, tools have been developed to allow QTL map alignments against consensus linkage maps, radiation hybrid (RH) maps, BAC fingerprinted contig (FPC) maps, single nucleotide polymorphism (SNP) location maps, and human maps. In addition, we have expanded the capabilities of the database such that research tools were developed where “private” preliminary QTL data could be entered and compared against all public data. This allows researchers to visualize data before publication and compare it with published results to aid in data interpretation. To serve this purpose, the database curator/editor tools also include functions that allow registered users to enter their own QTL data, make use of the QTLdb tools for data analysis, and use the QTLdb as a publishing tool (URL: ).