Mapping of genes expressed in activated porcine Peyer's patch

To determine the chromosomal locations for genes expressed in porcine Peyer's patches, polymerase chain reaction-based mapping of expressed sequence tags (ESTs) isolated from a porcine Peyer's patch-specific cDNA library was performed across a 6500-rad swine radiation hybrid panel. A total of 116 ESTs were mapped with LOD scores >6.0, and another 11 ESTs had LOD scores between 5.0 and 6.0. Of these 127 ESTs, 63% matched known genes (相似文献   

SNP detection and genetic mapping of porcine genes encoding enzymes in hepatic metabolic pathways and evaluation of linkage with carcass traits

We have previously identified and mapped porcine expressed sequence tags (ESTs) derived from genes that are preferentially expressed in liver. The aim of the present study was to identify single nucleotide polymorphisms (SNPs) in porcine genes encoding enzymes in hepatic metabolic pathways and use the SNPs for mapping. Furthermore, these genes, which are involved in utilization and partitioning of nutrients, were examined for their effects on carcass and meat quality traits by linkage analyses. In total, 100 ESTs were screened for SNPs by single strand conformation polymorphism analyses across a diverse panel of animals with a 36% success rate. Twelve of 36 polymorphic loci segregated in a three-generation Duroc x Berlin Miniature Pig (F2) resource population, the DUMI resource population, and were genetically mapped. Interval mapping of the corresponding chromosomes was performed to verify mapping of the genes within quantitative trait loci (QTL) regions detected in this resource population. QTL with genome-wide significance were detected in the vicinity of GNMT, ESTL147 and HGD. These loci therefore are positional candidate genes.     

Polymorphisms on SSC15q21-q26 Containing QTL for reproduction in Swine and its association with litter size

Several quantitative trait loci (QTL) for important reproductive traits (ovulation rate) have been identified on the porcine chromosome 15 (SSC15). To assist in the selection of positional candidate swine genes for these QTL on SSC15, twenty-one genes had already been assigned to SSC15 in a previous study in our lab, by using the radiation hybrid panel IMpRH. Further polymorphism studies were carried out on these positional candidate genes with four breeds of pigs (Duroc, Erhualian, Dahuabai and Landrace) harboring significant differences in reproduction traits. A total of nineteen polymorphisms were found in 21 genes. Among these, seven in six genes were used for association studies, whereby NRP2 polymorphism was found to be significantly (p < 0.05) associated with litter-size traits. NRP2 might be a candidate gene for pig-litter size based on its chromosome location (Du et al., 2006), significant association with litter-size traits and relationships with Sema and the VEGF super families.     

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.     

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.     

Comparative and physical mapping of 111 previously reported and 105 new porcine microsatellites

Here we report radiation hybrid mapping of 105 new porcine microsatellite markers on the IMpRH₇₀₀₀ radiation hybrid panel. In addition, we searched flanking sequences of these markers, as well as 673 previously reported RH-mapped microsatellite markers, for orthology to human sequences. Eighty-seven new and 111 previously mapped sequences exhibited orthology to human sequences. Using a stringent sequence alignment, 25 microsatellite-flanking sequences were found to be highly similar to genic sequences, whereas 173 were similar to non-genic sequences in the human genome. Five markers were located near known breakpoints of synteny between human and swine.     

Correlation between the presence of neutralizing antibodies against porcine circovirus 2 (PCV2) and protection against replication of the virus and development of PCV2-associated disease

Background

The rate of pubertal development and weaning to estrus interval are correlated and affect reproductive efficiency of swine. Quantitative trait loci (QTL) for age of puberty, nipple number and ovulation rate have been identified in Meishan crosses on pig chromosome 10q (SSC10) near the telomere, which is homologous to human chromosome 10p15 and contains an aldo-keto reductase (AKR) gene cluster with at least six family members. AKRs are tissue-specific hydroxysteroid dehydrogenases that interconvert weak steroid hormones to their more potent counterparts and regulate processes involved in development, homeostasis and reproduction. Because of their location in the swine genome and their implication in reproductive physiology, this gene cluster was characterized and evaluated for effects on reproductive traits in swine.

Results

Screening the porcine CHORI-242 BAC library with a full-length AKR1C4 cDNA identified 7 positive clones and sample sequencing of 5 BAC clones revealed 5 distinct AKR1C genes (AKR1CL2 and AKR1C1 through 4), which mapped to 126–128 cM on SSC10. Using the IMpRH_7000rad and IMNpRH2_12000rad radiation hybrid panels, these 5 genes mapped between microsatellite markers SWR67 and SW2067. Comparison of sequence data with the porcine BAC fingerprint map show that the cluster of genes resides in a 300 kb region. Twelve SNPs were genotyped in gilts observed for age at first estrus and ovulation rate from the F8 and F10 generations of one-quarter Meishan descendants of the USMARC resource population. Age at puberty, nipple number and ovulation rate data were analyzed for association with genotypes by MTDFREML using an animal model. One SNP, a phenylalanine to isoleucine substitution in AKR1C2, was associated with age of puberty (p = 0.07) and possibly ovulation rate (p = 0.102). Two SNP in AKR1C4 were significantly associated with nipple number (p ≤ 0.03) and another possibly associated with age at puberty (p = 0.09).

Conclusion

AKR1C genotypes were associated with nipple number as well as possible effects on age at puberty and ovulation rate. The estimated effects of AKR1C genotypes on these traits suggest that the SNPs are in incomplete linkage disequilibrium with the causal mutations that affect reproductive traits in swine. Further investigations are necessary to identify these mutations and understand how these AKR1C genes affect these important reproductive traits. The nucleotide sequence data reported have been submitted to GenBank and assigned accession numbers [GenBank:DQ474064–DQ474068, GenBank:DQ494488–DQ494490 and GenBank:DQ487182–DQ487184].     

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 in the pig: localization of 214 expressed sequence tags

In total, 214 ESTs (Expressed Sequence Tags) were assigned to the porcine gene map by using somatic cell hybrid mapping, radiation hybrid mapping, and FISH. The ESTs were isolated from a porcine small intestine cDNA library on the basis of significant sequence identity with human annotated genes. In total, 390 primer pairs were designed primarily in the 3' UTR of the sequences. Overall, 58.6% of the ESTs were successfully mapped by this approach. In total, 191 of the localizations are in agreement with the human comparative map, strongly indicating that these represent true orthologous genes. The remaining 23 ESTs provide new comparative mapping data, which should be considered as preliminary until confirmed by other studies. Our mapping efforts provide a significant contribution to the porcine map as well as to the comparative map for human and pig.     

Mapping and expression analyses during porcine foetal muscle development of 12 genes involved in histone modifications

Histone modifications (methylation and demethylation) regulate gene expression and play a role in cell proliferation and differentiation by their actions on chromatin structure. In this context, we studied the temporal expression profiles of genes acting on histone methylation and demethylation during skeletal muscle proliferation and differentiation. Quantitative real-time PCR was used to quantify the mRNA levels of CARM1 , JARID1A , JMJD2A , LSD1 , PRMT2 , PRMT5 , SMYD1 , SMYD2 , SMYD3 , SETDB1 , Suv39h2 and SUZ12 in foetal skeletal muscle. Our results showed that CARM1 , JARID1A , JMJD2A , SMYD1 and SMYD2 were differentially expressed in embryonic muscles of 33 days post-conception (dpc), 65 dpc and 90 dpc. These 12 genes were mapped to porcine chromosomes (SSC) 2q21–24, 5q25, 6q35, 6q12–21, 6p15, 7q21, 3q21–27, 9q26, 10p16, 4q15–16, 10q14–16 and 12p12 respectively. Taking into account the reported QTL mapping results, gene expression analysis and radiation hybrid mapping results, these results suggest that five genes ( CARM1 , JARID1A , JMJD2A , SMYD1 and SMYD2 ) could be good candidate genes for growth and backfat thickness traits.     

Comparative analysis and development of microsatellite markers on swine (Sus scrofa) chromosome 1qter

Several quantitative trait loci (QTL) have been detected on SSC1qter (Sus scrofa chromosome 1qter), including QTL for the number of vertebrae, as reported in our previous study. To provide the tools for analysis of QTLs on SSC1qter, we constructed a comparative map of swine and human. In addition, we identified 26 swine STSs and mapped 16 of them on SSC1qter using the INRA - University of Minnesota porcine radiation hybrid (IMpRH) panel. We screened a BAC library using these swine STSs and developed 35 new polymorphic microsatellite markers from the BAC clones, of which 26 were informative in our reference family. We also mapped nine microsatellite markers we had isolated previously. Consequently a total of 44 new polymorphic microsatellite markers were located within a 60-cM region of SSC1qter, spanning from SW1092 to the telomere.     

Linking porcine microsatellite markers to known genome regions by identifying their human orthologs.

Microsatellites, or tandem simple sequence repeats (SSRs), have become one of the most popular molecular markers in genome mapping because of their abundance across genomes and because of their high levels of polymorphism. However, information on which genes surround or flank them has remained very limited for most SSRs, especially in livestock species. In this study, an in silico comparative mapping approach was developed to link porcine SSRs to known genome regions by identifying their human orthologs. From a total of 1321 porcine microsatellites used in this study, 228 were found to have blocks in alignment with human genomic sequences. These 228 SSRs span about 1459 cM of the porcine genome, but with uneven distributions, ranging from 2 on SSC12 to 24 on SSC14. Linking these porcine SSRs to the known genome regions in the human genome also revealed 16 new putative synteny groups between these two species. Fifteen SSRs on SSC3 with identified human orthologs were typed on a pig-hamster radiation hybrid (RH) panel and used in a joint analysis with 80 known gene markers previously mapped on SSC3 using the same panel. The analysis revealed that they were all highly linked to either one or both adjacent markers. These results indicated that assigning the porcine SSRs to known genome regions by identifying their human orthologs is a reliable approach. The process will provide a foundation for positional cloning of causative genes for economically important traits.     

Comparative mapping of RPL3, a gene overexpressed in multiple obesity models.

The ribosomal protein 3 gene is differentially expressed in hypothalamus and brown adipose tissue between mouse lines divergently selected for heat loss, and in skeletal muscle of the ob/ob mouse model. Unfortunately, multiple Rpl3-processed pseudogenes have hampered mapping of the functional gene copy in mammalian species. Using PCR amplification with intronic primer binding, we have mapped Rpl3 to MMU15, and have also localized RPL3 to BTA5 in cattle. Comparative mapping implicates a previously mapped copy of RPL3 on HSA22 as the functional copy of human RPL3, while predictive mapping places the porcine homologue on SSC5.     

Chromosomal localization of ESTs obtained from human fetal liver via BAC-mediated FISH mapping.

A total of 55 expressed sequence tags (ESTs) randomly chosen from our collection of fetal liver ESTs were mapped to chromosomes by fluorescence in situ hybridization (FISH) mapping techniques. To generate FISH mapping probes, the genomic DNAs for each EST were selected by screening an arrayed human bacterial artificial chromosome (BAC) library. In total, 73 BACs were used for mapping of the 55 ESTs. Among them, 70 BACs representing 52 ESTs unequivocally mapped to single chromosomal regions. The remaining 3 BACs representing 3 ESTs were localized to multiple regions, suggesting that BACs may have very low chimerism. Our mapping results were compared with EST mapping databases deposited in NCBI. Thirty-six of 55 ESTs corresponded to previously mapped positions of ESTs, 2 ESTs mapped to different positions from previously determined ones, and it was found that 17 ESTs have been mapped on new locations from this study. These mapping data may be used for completing the framework of the human physical map, and also for providing a good starting point for searching disease-related genes.