Zebrafish Genetic Map with 2000 Microsatellite Markers

The zebrafish is the first vertebrate organism used for large-scale genetic screens seeking genes critical to development. These screens have been quite successful, with more than 1800 recessive mutations discovered that speak to morphogenesis of the vertebrate embryo. The cloning of the mutant genes depends on a dense genetic map. The 2000 markers we present here, using microsatellite (CA) repeats, provides 1.2-cM average resolution. One centimorgan in zebrafish is about 0. 74 megabase, so, for many mutations, these markers are close enough to begin positional cloning by YAC walks.     

Integration of the Physical and Genetic Linkage Map for Human Chromosome 13

We have used a panel of somatic cell hybrids containing different rearrangements of human chromosome 13 to integrate genetic and physical maps of this chromosome. The positions of 17 translocation/deletion breakpoints on human chromosome 13 have been determined relative to the microsatellite markers on the genetic linkage map compiled by Généthon. Because markers on maps from several other Consortium groups have also been analyzed using many of the same hybrids, it was possible to relate these with the Généthon map. The position of all of the chromosome breakpoints have been placed, wherever possible, between two adjacent markers on the genetic linkage maps using PCR analysis for the presence/absence of the markers in the somatic cell hybrids. The positions of the breakpoints have already been determined cytogenetically, and some of these breakpoints are located at landmark positions on the chromosome. The relative density of markers along the chromosome differs between independently derived maps, and, based on the known locations of certain breakpoints in the physical map, inconsistencies in the genetic maps have been identified.     

Physical Mapping of Genetic Markers on the Short Arm of Chromosome 5

The deletion of the short arm of chromosome 5 is associated with the cri-du-chat syndrome. In addition, loss of this portion of a chromosome is a common cytogenetic marker in a number of malignancies. However, to date, no genes associated with these disorders have been identified. Physical maps are the first step in isolating causative genes, and genes involved in autosomal recessive disorders are now routinely mapped through the identification of linked markers. Extensive genetic maps based upon polymorphic short tandem repeats (STRs) have provided researchers with a large number of markers to which such disorders can be genetically mapped. However, the physical locations of many of these STRs have not been determined. Toward the goal of integrating the human genetic maps with the physical maps, a 5p somatic cell hybrid deletion mapping panel that was derived from patients with 5p deletions or translocations was used to physically map 47 STRs that have been used to construct genetic maps of 5p. These data will be useful in the localization of disease genes that map to 5p and may be involved in the etiology of the cri-du-chat syndrome.     

微卫星标记对资源猪群的遗传分析和连锁图谱构建

在猪数量性状位点的定位研究中,标记的使用和图谱的构建是很重要的。本研究从猪的第4、6、7、8和13染色体上选取39个微卫星标记,在来源于约克夏和梅山214头猪组成的资源群中,分析了遗传特征并构建了图谱。研究表明,平均等位基因数、平均观察杂合度(Ho)和平均多态信息含量(PIC)在F1和F2代中分别为:3.2,0.528,0.463和3.2,0.496,0.447。结果表明大多数微卫星标记位点表现为中高度杂合性。在资源群体中,平均有信息减数分裂数是217.4(44-316),而各染色体上两性平均图谱的长度分别是:172.3cM(SSC4),168.7cM(SSC6),191.7cM(SSC7),197.3cM(SSC8),178.3cM(SSC13)。与USDA-MARC的参考图谱相比,标记位点的顺序相同,但长度均较长。雌雄两性图谱相比,第4和第6染色体上雌性图谱长于雄性图谱;而在另外3条染色体上,则雄性图谱长于雌性图谱。结果显示了标记位点在资源猪群的遗传特征和遗传关系,其连锁图谱可用于今后的QTL定位。     

Anchoring Linkage Groups of the Rosa Genetic Map to Physical Chromosomes with Tyramide-FISH and EST-SNP Markers

In order to anchor Rosa linkage groups to physical chromosomes, a combination of the Tyramide-FISH technology and the modern molecular marker system based on High Resolution Melting (HRM) is an efficient approach. Although, Tyramide-FISH is a very promising technique for the visualization of short DNA probes, it is very challenging for plant species with small chromosomes such as Rosa. In this study, we successfully applied the Tyramide-FISH technique for Rosa and compared different detection systems. An indirect detection system exploiting biotinylated tyramides was shown to be the most suitable technique for reliable signal detection. Three gene fragments with a size of 1100 pb–1700 bp (Phenylalanine Ammonia Lyase, Pyrroline-5-Carboxylate Synthase and Orcinol O-Methyl Transferase) have been physically mapped on chromosomes 7, 4 and 1, respectively, of Rosa wichurana. The signal frequency was between 25% and 40%. HRM markers of these 3 gene fragments were used to include the gene fragments on the existing genetic linkage map of Rosa wichurana. As a result, three linkage groups could be anchored to their physical chromosomes. The information was used to check for synteny between the Rosa chromosomes and Fragaria.     

Dinucleotide Repeat Loci Contribute Highly Informative Genetic Markers to the Human Chromosome 2 Linkage Map

Microsatellite repeat loci can provide informative markers for genetic linkage. Currently, the human chromosome 2 genetic linkage map has very few highly polymorphic markers. Being such a large chromosome, it will require a large number of informative markers for the dense coverage desired to allow disease genes to be mapped quickly and accurately. Dinucleotide repeat loci from two anonymous chromosome 2 genomic DNA clones were sequenced so that oligonucleotide primers could be designed for amplifying each locus using the polymerase chain reaction (PCR). Five sets of PCR primers were also generated from nucleotide sequences in the GenBank Database of chromosome 2 genes containing dinucleotide repeats. In addition, one PCR primer pair was made that amplifies a restriction fragment length polymorphism on the TNP1 gene (Hoth and Engel, 1991). These markers were placed on the CEPH genetic linkage map by screening the CEPH reference DNA panel with each primer set, combining these data with those of other markers previously placed on the map, and analyzing the combined data set using CRI-MAP and LINKAGE. The microsatellite loci are highly informative markers and the TNP1 locus, as expected, is only moderately informative. A map was constructed with 38 ordered loci (odds 1000:1) spanning 296 cM (male) and 476 cM (female) of chromosome 2 compared with 306 cM (male) and 529 cM (female) for a previous map of 20 markers.     

In Silico Identification and Mapping of Microsatellite Markers on Sus Scrofa Chromosome 4

Marker density of a QTL region on pig chromosome 4 was increased. New microsatellites were identified by in silico mining of BAC-end and genomic shotgun sequences. Among 8,784 BAC-end sequences predicted within the region, 148 microsatellites were identified. In addition, 27,450 CA/TG repeats were identified within the genomic shotgun sequences, of which 157 were most likely located on SSC4q. A selection of 61 new microsatellites was mapped, together with previously mapped markers. The results showed that the human-pig comparative map in combination with BAC-end and genomic sequence resources provides an excellent source for a highly efficient and targeted development of markers.     

Genetic Linkage Mapping of the Dehydroepiandrosterone Sulfotransferase (STD) Gene on the Chromosome 19q13.3 Region

In the human liver and adrenal, there is a single hydroxysteroid sulfotransferase, which catalyzes the transformation of dehydroepiandrosterone to dehydroepiandrosterone sulfate, the most abundantly circulating steroid in humans, and also catalyzes the sulfation of a series of other 3β-hydroxysteroids as well as cholesterol. Dehydroepiandrosterone sulfate serves as precursor for the formation of active androgens and estrogens in several peripheral tissues, indicating that hydroxysteroid sulfotransferase plays a pivotal role in controlling the hormonal action of sex steroids by regulating their bioavailability. We recently elucidated the structure of the gene encoding hydroxysteroid sulfotransferase (STD), also designated dehydroepiandrosterone sulfotransferase, which spans 17 kb and contains six exons. The STD gene was preliminarily assigned to chromosome 19 by polymerase chain reaction (PCR) amplification of DNA from a panel of human/rodent somatic cell hybrids. To locate the STD gene, the novel biallelic polymorphism found in intron 2 was genotyped in eight CEPH reference families by direct sequencing of PCR products. Two-point linkage analysis was first performed between the latter polymorphism and chromosome 19 markers from Généthon and NIH/CEPH. The closest linkage was observed with D19S412 (Z_max= 9.23; θ_max0.038) and HRC (Z_max= 5.95; θ_max0.036), located on the 19q13.3 region. A framework map including six Généthon markers flanking the polymorphic STD gene was created by multipoint linkage analysis. Thereafter, a high-resolution genetic map of the region was constructed, yielding to the following order: qter–D19S414–D19S224–D19S420–D19S217–(APOC2–D19S412)–(STD–HRC)– KLK–D19S22–D19S180–PRKCG–D19S418–tel.     

猪1号染色体微卫星多态性研究及遗传连锁图谱的构建

微卫星具有多态性高、保守性好等优点。本研究选取以20cM左右的间距均匀分布于1号染色体的8个多态微卫星基因座构建猪1号染色体的遗传连锁图谱,为进一步进行重要经济性状基因座的定位打下基础。试验结果表明,8个基因座等位基因数目2~5个,各个基因座等位基因频率在0.015~0.75之间,杂合度为0.39705~0.67675,多态信息含量为0.32925~0.59316。构建的资源家系遗传连锁图谱总长181.5cM,与USDA结果基本一致,可进一步用于猪数量性状基因座定位的研究。 The Microsatellite Polymorphism Research on Porcine Chromosome 1 and the Construction of Its Genetic Map QU Yan-chun,DENG Chang-yan,XIONG Yuan-zhu,SU Yu-hong,ZHENG Rong,LIU Gui-lan The Key Laboratory of Pig Breeding and Genetics,The Ministry of Agriculture, Huazhong Agricultural University,Wuhan 430070,China Abstract:As a molecular marker,microsatellite has many advantages such as high polymorphism and good conservativeness in animal genetic research.The study chose 8 microsatellite markers that evenly distributed on chromosome 1 with a distance about 20 cM to build the genetic map of porcine chromosome 1.The results of our experiment are as follows:the number of alleles for 8 markers is 2 to 5,their gene frequency is from 0.015 to 0.75,the heterozygosity is from 0.39705 to 0.67675 and the polymorphic information content is from 0.32925 to 0.59316.The map we built is basically in consistent with the result of USDA and can be used in searching quantitative traits loci in pigs. Key words：porcine; microsatellite; heterozygosity; polymorphic information content; genetic map     

Genetic and Physical Mapping of the Dreher Locus on Mouse Chromosome 1

Mutations in the mouse dreher (dr) gene cause skeletal defects, hyperactivity, abnormal gait, deafness, white belly spotting, and hypoplasia of Müllerian duct derivatives. To map dr to high resolution, we utilized two crosses. Initially, we analyzed an intersubspecific intercross to construct a detailed genetic map of simple sequence length polymorphism markers within a 6.3-cM region surrounding the dr locus. Subsequently, we analyzed a second intersubspecific intercross segregating for the dr(6J) allele, which positioned dr within a 0.13-cM region between Rxrg and D1Mit370. A physical contig of BAC clones spanning the dr critical region was constructed, and eight potential dr candidate genes were excluded by genetic or physical mapping. Together these results lay the foundation for positional cloning of the dr gene.     

用商品群作为参考系构建猪的微卫星连锁图谱

由 19头杂种公猪 [皮特兰× (皮特兰×汉普夏 ) ]、5 2头杂种母猪 [Leicoma× (大约克×长白 ) ]及其 332头后代组成的商品群作为参考系 ,选择 172个微卫星标记和 3个Ⅰ类标记 (RYR1、PIT1、PRKAG3)对参考系的个体进行遗传标记分型 ,应用CRIMAP(2 4 )构建猪的整个基因组微卫星连锁图谱。采用多重PCR方法对微卫星标记进行扩增 ,用ABI 377测序仪进行电泳分离 ,应用Genescan 3 0和Genotyper 2 0软件进行基因型检测。 3个Ⅰ类标记用PCR RFLP技术进行分型。CRIMAP程序分析表明 :所构建的猪常染色体性平均连锁图谱的总长度为 2 36 8 7cM ,X染色体的长度为 14 3 10cM ,遗传标记的平均间距为 16 3cM ,亲本的微卫星标记座位的杂合度平均为 0 70。此连锁图谱的构建将为商品猪群的生长、胴体、肉质以及繁殖性状的QTL扫描打下基础。     

A Genetic Linkage Map of Mouse Chromosome 10: Localization of Eighteen Molecular Markers Using a Single Interspecific Backcross

Interspecific mouse backcross analysis was used to generate a molecular genetic linkage map of mouse chromosome 10. The map locations of the Act-2, Ahi-1, Bcr, Braf, Cdc-2a, Col6a-1, Col6a-2, Cos-1, Esr, Fyn, Gli, Ifg, Igf-1, Myb, Pah, pgcha, Ros-1 and S100b loci were determined. These loci extend over 80% of the genetic length of the chromosome, providing molecular access to many regions of chromosome 10 for the first time. The locations of the genes mapped in this study extend the known regions of synteny between mouse chromosome 10 and human chromosomes 6, 10, 12 and 21, and reveal a novel homology segment between mouse chromosome 10 and human chromosome 22. Several loci may lie close to, or correspond to, known mutations. Preferential transmission of Mus spretus-derived alleles was observed for loci mapping to the central region of mouse chromosome 10.     

A Microsatellite Linkage Map of the Porcine Genome

We report the most extensive genetic linkage map for a livestock species produced to date. We have linked 376 microsatellite (MS) loci with seven restriction fragment length polymorphic loci in a backcross reference population. The 383 markers were placed into 24 linkage groups which span 1997 cM. Seven additional MS did not fall into a linkage group. Linkage groups are assigned to 13 autosomes and the X chromosome (haploid n = 19). This map provides the basis for genetic analysis of quantitative inheritance of phenotypic and physiologic traits in swine.     

Physical and Genetic Map of the Pasteurella multocida A:1 Chromosome

A physical and genetic map of the Pasteurella multocida A:1 genome was generated by using the restriction enzymes ApaI, CeuI, and NotI. The positions of 23 restriction sites and 32 genes, including 5 rrn operons, were localized on the 2.35-Mbp single circular chromosome. This report presents the first genetic and physical map for this genus.     

Combined Physical and Genetic Map of the Pseudomonas putida KT2440 Chromosome

A combined physical and genetic map of the Pseudomonas putida KT2440 genome was constructed from data obtained by pulsed-field gel electrophoresis techniques (PFGE) and Southern hybridization. Circular genome size was estimated at 6.0 Mb by adding the sizes of 19 SwaI, 9 PmeI, 6 PacI, and 6 I-CeuI fragments. A complete physical map was achieved by combining the results of (i) analysis of PFGE of the DNA fragments resulting from digestion of the whole genome with PmeI, SwaI, I-CeuI, and PacI as well as double digestion with combinations of these enzymes and (ii) Southern hybridization analysis of the whole wild-type genome digested with different enzymes and hybridized against a series of probes obtained as cloned genes from different pseudomonads of rRNA group I and Escherichia coli, as P. putida DNA obtained by PCR amplification based on sequences deposited at the GenBank database, and by labeling of macrorestriction fragments of the P. putida genome eluted from agarose gels. As an alternative, 10 random mini-Tn5-Km mutants of P. putida KT2440 were used as a source of DNA, and the band carrying the mini-Tn5 in each mutant was identified after PFGE of a series of complete chromosomal digestions and hybridization with the kanamycin resistance gene of the mini-Tn5 as a probe. We established a circular genome map with an average resolution of 160 kb. Among the 63 genes located on the genetic map were key markers such as oriC, 6 rrn loci (rnnA to -F), recA, ftsZ, rpoS, rpoD, rpoN, and gyrB; auxotrophic markers; and catabolic genes for the metabolism of aromatic compounds. The genetic map of P. putida KT2440 was compared to those of Pseudomonas aeruginosa PAO1 and Pseudomonas fluorescens SBW25. The chromosomal backbone revealed some similarity in gene clustering among the three pseudomonads but differences in physical organization, probably as a result of intraspecific rearrangements.     

Development of Polymorphic EST Markers Suitable for Genetic Linkage Mapping of Catfish

Expressed sequence tag (EST) markers are important for gene mapping and for marker-assisted selection (MAS). To develop EST markers for use in catfish gene mapping, 100 randomly picked complementary DNAs from the channel catfish (Ictalurus punctatus) pituitary library were sequenced. The EST sequences were used to design primers to amplify channel catfish and blue catfish (I. furcatus) genomic DNAs. Polymerase chain reaction products of the ESTs were analyzed to determine length polymorphism between the channel catfish and blue catfish. Eleven polymorphic EST markers were identified. Five of the 11 EST markers were from known genes and the other six were from unidentified ESTs. Seven ESTs were found to be associated with microsatellite sequences. Analysis of channel catfish gene sequences indicated highly biased codon usage, with 16 codons being preferably used. These codons were more preferably used in highly expressed ribosomal protein genes and in highly expressed pituitary hormone genes. G/C-rich codons are less used in channel catfish than those in other vertebrates suggesting AT-richness of the channel catfish genome. Received June 29, 1998; accepted March 29, 1999.     

Construction of a High-Density Microsatellite Genetic Linkage Map and Mapping of Sexual and Growth-Related Traits in Half-Smooth Tongue Sole (Cynoglossus semilaevis)

High-density genetic linkage maps of half-smooth tongue sole were developed with 1007 microsatellite markers, two SCAR markers and an F1 family containing 94. The female map was composed of 828 markers in 21 linkage groups, covering a total of 1447.3 cM, with an average interval 1.83 cM between markers. The male map consisted of 794 markers in 21 linkage groups, spanning 1497.5 cM, with an average interval of 1.96 cM. The female and male maps had 812 and 785 unique positions, respectively. The genome length of half-smooth tongue sole was estimated to be 1527.7 cM for the females and 1582.1 cM for the males. Based on estimations of the map lengths, the female and male maps covered 94.74 and 94.65% of the genome, respectively. The consensus map was composed of 1007 microsatellite markers and two SCAR markers in 21 linkage groups, covering a total of 1624 cM with an average interval of 1.67 cM. Furthermore, 159 sex-linked SSR markers were identified. Five sex-linked microsatellite markers were confirmed in their association with sex in a large number of individuals selected from different families. These sex-linked markers were mapped on the female map LG1f with zero recombination. Two QTLs that were identified for body weight, designated as We-1 and We-2, accounted for 26.39% and 10.60% of the phenotypic variation. Two QTLs for body width, designated Wi-1 and Wi-2, were mapped in LG4f and accounted for 14.33% and 12.83% of the phenotypic variation, respectively. Seven sex-related loci were mapped in LG1f, LG14f and LG1m by CIM, accounting for 12.5–25.2% of the trait variation. The results should prove to be very useful for improving growth traits using molecular MAS.     

RAPD和SSR两种标记构建的中国对虾遗传连锁图谱

利用RAPD和SSR分子标记结合拟测交策略,对中国对虾(Fenneropenaeuschinensis)“黄海1号”雌虾与野生雄虾作为亲本进行单对杂交产生的F1代,采用RAPD和SSR两种分子标记技术初步构建了中国对虾雌、雄遗传连锁图谱。对460个RAPD引物和44对SSR引物进行筛选,共选出61个RAPD引物和20对SSR引物,用于对父母本和82个F1个体进行遗传分析。共得到母本分离标记146个(RAPD标记128个,微卫星标记18个)和父本分离标记127个(RAPD标记109个,微卫星标记18个)。雌性图谱包括8个连锁群、9个三联体和14个连锁对,标记间平均间隔为11·28cM,图谱共覆盖1173cM,覆盖率为59·36%;雄性图谱包括10个连锁群、12个三联体和7个连锁对,标记间平均间隔为12·05cM,图谱共覆盖1144·6cM,覆盖率为62·01%。中国对虾遗传图谱的构建为其分子标记辅助育种、比较基因组作图及数量性状位点的定位与克隆奠定了基础。     

Genomic Characterization of DArT Markers Based on High-Density Linkage Analysis and Physical Mapping to the Eucalyptus Genome

Diversity Arrays Technology (DArT) provides a robust, high throughput, cost-effective method to query thousands of sequence polymorphisms in a single assay. Despite the extensive use of this genotyping platform for numerous plant species, little is known regarding the sequence attributes and genome-wide distribution of DArT markers. We investigated the genomic properties of the 7,680 DArT marker probes of a Eucalyptus array, by sequencing them, constructing a high density linkage map and carrying out detailed physical mapping analyses to the Eucalyptus grandis reference genome. A consensus linkage map with 2,274 DArT markers anchored to 210 microsatellites and a framework map, with improved support for ordering, displayed extensive collinearity with the genome sequence. Only 1.4 Mbp of the 75 Mbp of still unplaced scaffold sequence was captured by 45 linkage mapped but physically unaligned markers to the 11 main Eucalyptus pseudochromosomes, providing compelling evidence for the quality and completeness of the current Eucalyptus genome assembly. A highly significant correspondence was found between the locations of DArT markers and predicted gene models, while most of the 89 DArT probes unaligned to the genome correspond to sequences likely absent in E. grandis, consistent with the pan-genomic feature of this multi-Eucalyptus species DArT array. These comprehensive linkage-to-physical mapping analyses provide novel data regarding the genomic attributes of DArT markers in plant genomes in general and for Eucalyptus in particular. DArT markers preferentially target the gene space and display a largely homogeneous distribution across the genome, thereby providing superb coverage for mapping and genome-wide applications in breeding and diversity studies. Data reported on these ubiquitous properties of DArT markers will be particularly valuable to researchers working on less-studied crop species who already count on DArT genotyping arrays but for which no reference genome is yet available to allow such detailed characterization.