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Biomedical subjects

G A Rohrer

Publications and source records attributed to G A Rohrer.

At least 37 records · Page 2Linked to original sources

Molecular cloning of the porcine inhibin-betaB gene and reassignment to chromosome 15.

Inhibins are gonadal glycoproteins belonging to the transforming growth factor-beta superfamily that act to suppress pituitary follicle stimulating hormone and are composed of a common alpha-subunit linked by disulphide bonds to either a betaA- or betaB-subunit. The porcine inhibin-alpha, -betaA (INHBA) and -betaB (INHBB) subunit genes have previously been mapped to chromosomes 15, 18 and 12, respectively. Over 6.7 kb of the INHBB gene was sequenced from a porcine genomic cosmid clone and found to contain two microsatellites, one in intron 1 and the other in the 3'-untranslated region. Both microsatellites mapped to pig chromosome 15 at relative position 48 cm. This sequence was greater than 99% identical to two previously reported partial non-contiguous cDNAs for porcine INHBB. Non-coding regions also had a high degree (79-88%) of identity with the corresponding regions of the human gene. Based on sequence information and mapping of two novel microsatellite markers, we reassigned porcine INHBB to chromosome 15, which is consistent with comparative physical and linkage maps of this chromosome and human chromosome 2.

Animals↗

Relationships of testicular iron and ferritin concentrations with testicular weight and sperm production in boars.

The inverse relationship of testicular size and circulating follicle-stimulating hormone (FSH) concentrations has been documented, and accompanying this relationship is the change in color of the parenchymal tissue of the testes. Large testes (300 to 400 g) are pink to light red and small testes (100 g) are dark maroon with color gradations for weights in between. It was hypothesized that this color most likely represented an iron protein. Chromatographic analysis of testicular tissue indicated that the Fe was associated primarily with ferritin, and immunohistochemistry showed that Leydig cells were the primary location of ferritin storage within the testes. Concentrations of Fe and ferritin were higher in small testes and decreased as testes weight increased (P < 0.05). As testicular Fe concentrations increased, daily sperm production (DSP) and total DSP declined (P < 0.05). Genotyping six generations of Meishan x White composite boars (n = 288) for a quantitative trait locus that is indicative of elevated FSH and small testes in boars indicated that the Meishan genotype had elevated testicular iron concentrations and darker color in conjunction with reduced total DSP (P < 0.01). It is not thought the elevated iron concentrations affect testicular weights but are probably a result of elevated FSH and FSH inducement of Fe transport. The storage of Fe in Leydig cells may provide a reservoir of Fe for easy access by Sertoli and germ cells, but still provide a degree of protection to germ cells from ionic iron.

Animals↗

Fine mapping a quantitative trait locus affecting ovulation rate in swine on chromosome 8.

Ovulation rate is an integral component of litter size in swine, but is difficult to directly select for in commercial swine production. Because a QTL has been detected for ovulation rate at the terminal end of chromosome 8p, genetic markers for this QTL would enable direct selection for ovulation rate in both males and females. Eleven genes from human chromosome 4p16-p15, as well as one physiological candidate gene, were genetically mapped in the pig. Large insert swine genomic libraries were screened, clones were isolated and then screened for microsatellite repeats, and informative microsatellite markers were developed for seven genes (GNRHR, IDUA, MAN2B2, MSX1, PDE6B, PPP2R2C, and RGS12). Three genes (LRPAP1, GPRK2L, and FLJ20425) were mapped using genotyping assays developed from single nucleotide polymorphisms. Two genes were assigned since they were present in clones that contained mapped markers (HGFAC and HMX1). The resulting linkage map of pig chromosome 8 contains markers associated with 14 genes in the first 27 cM. One inversion spanning at least 3 Mb in the human genome was detected; all other differences could be explained by resolution of mapping techniques used. Fourteen of the most informative microsatellite markers in the first 27 cM of the map were genotyped across the entire MARC swine resource population, increasing the number of markers typed from 2 to 14 and more than doubling the number ofgenotyped animals with ovulation rate data (295 to 600). Results from the revised data set for the QTL analysis, assuming breed specific QTL alleles, indicated that the most likely position of the QTL resided at 4.85 cM on the new linkage map (F1,592 = 20.5150, genome-wide probability less than 0.015). The updated estimate of the effect of an allele substitution was -1.65 ova for the Meishan allele. The F-ratio peak was closest to markers for MAN2B2 (4.80 cM) and was flanked on the other side by markers for PPP2R2C. Two positional candidate genes included in this study are MAN2B2 and RGS12. These results validate the presence of a QTL affecting ovulation rate on chromosome 8 and facilitate selection of positional candidate genes to be evaluated.

Animals↗

Characterization of porcine uterine estrogen sulfotransferase.

A quantitative trait locus (QTL) for uterine capacity is located on chromosome 8. Comparison of porcine and human genetic maps suggested that the estrogen sulfotransferase (STE) gene may be located near this region. The objectives of this study were to clone the full coding region for STE, compare endometrial STE gene expression between Meishan and White composite pigs during early pregnancy, and map the STE gene. We obtained a clone (1886 bp) containing the full coding region of STE by iterative screening of an expressed sequence tag library. Endometrial STE mRNA expression in White composite gilts was determined by Northern blotting on days 10, 13, and 15 of the estrous cycle; and on days 10, 13, 15, 20, 30, and 40 of pregnancy. STE mRNA expression was elevated (P < 0.01) on days 20 and 30 of pregnancy compared to other days of the cycle or pregnancy. Endometrial STE mRNA expression during early pregnancy, determined using real-time RT-PCR, was elevated (P < 0.01) on day 20 compared to day 15, decreased (P = 0.02) between days 20 and 30, and decreased further (P < 0.01) between days 30 and 40 in both Meishan and White composite pigs. Expression of STE mRNA was greater (P = 0.01) in White composite pigs compared to Meishan pigs. Using a microsatellite from an STE containing BAC genomic clone, the STE gene was mapped to 65 centimorgans on chromosome 8. Because STE mRNA expression differs between Meishan and White composite pigs, the STE gene may be a candidate for the uterine capacity QTL.

Amino Acid Sequence↗

Single nucleotide polymorphism (SNP) discovery in porcine expressed genes.

High-throughput genotyping of swine populations is a potentially efficient method for establishing animal lineage and identification of loci important to animal health and efficient pork production. Markers were developed based upon single nucleotide polymorphisms (SNPs), which are abundant and amenable to automated genotyping platforms. The focus of this research was SNP discovery in expressed porcine genes providing markers to develop the porcine/human comparative map. Locus specific amplification (LSA) and comparative sequencing were used to generate PCR products and allelic information from parents of a swine reference family. Discovery of 1650 SNPs in 403 amplicons and strategies for optimizing LSA-based SNP discovery using alternative methods of PCR primer design, data analysis, and germplasm selection that are applicable to other populations and species are described. These data were the first large-scale assessment of frequency and distribution of porcine SNPs.

Animals↗

An integrated comparative map of the porcine X chromosome.

The objectives of this study were to assign both microsatellite and gene-based markers on porcine chromosome X to two radiation hybrid (RH) panels and to develop a more extensive integrated map of SSC-X. Thirty-five microsatellite and 20 gene-based markers were assigned to T43RH, and 16 previously unreported microsatellite and 15 gene-based markers were added to IMpRH map. Of these, 30 microsatellite and 12 gene-based markers were common to both RH maps. Twenty-two gene-based markers were submitted to BLASTN analysis for identification of orthologues of genes on HSA-X. Single nucleotide polymorphisms (SNPs) were detected for 12 gene-based markers, and nine of these were placed on the genetic map. A total of 92 known loci are present on at least one porcine chromosome X map. Thirty-seven loci are present on all three maps; 31 loci are found on only one map. Location of 33 gene-based markers on the comprehensive map translates into an integrated comparative map that supports conservation of gene order between SSC-X and HSA-X. This integrated map will be valuable for selection of candidate genes for porcine quantitative trait loci (QTLs) that map to SSC-X.

Animals↗

Mapping microsatellite markers identified in porcine EST sequences.

A sequence search of swine expressed sequence tags (EST) data in GenBank identified over 100 sequence files which contained a microsatellite repeat or simple sequence repeat (SSR). Most of these repeat motifs were dinucleotide (CA/GT) repeats; however, a number of tri-, tetra-, penta- and hexa-nucleotide repeats were also detected. An initial assessment of six dinucleotide and 14 higher-order repeat markers indicated that only dinucleotide markers yielded a sufficient number of informative markers (100% vs. 14% for dinucleotide and higher order repeats, respectively). Primers were designed for an additional 50 di- and one tri-nucleotide SSRs. Overall, 42 markers were polymorphic in the US Meat Animal Research Center (MARC) reference population, 17 markers were uninformative and 12 primer pairs failed to satisfactorily amplify genomic DNA. A comparison of di-nucleotide repeat vs. markers with repeat motifs of three to six bases demonstrated that 72% of dinucleotide markers were informative relative to only 7% of other repeat motifs. The difference was the result of a much higher percentage of monomorphic markers in the three to six base repeat motif markers than in the dinucleotide markers (64% vs. 14%). Either higher order repeat motifs are less polymorphic in the porcine genome or our selection criteria for repeat length of more than 17 contiguous bases was too low. The mapped microsatellite markers add to the porcine genetic map and provide valuable links between the porcine and human genome.

Alleles↗

Characterization and chromosome assignment of the porcine AHCY gene for S-adenosylhomocysteine hydrolase.

The gene for S-adenosylhomocysteine hydrolase (AHCY) is involved in the regulation of cellular methylation reactions. Here we report the cloning, sequencing, and chromosomal assignment of the porcine AHCY gene. The gene consists of 10 exons spanning approximately 19 kb of genomic DNA. It encodes a protein of 432 amino acids that shows about 96% identity to the orthologous S-adenosylhomocysteine hydrolases from human, rat, and mouse. The porcine AHCY gene is located very close to the agouti signaling protein gene (ASIP) on SSC17q21. The chromosomal localization was subsequently confirmed by RH mapping and the genetic mapping of an intragenic microsatellite that maps to 57 cM on the linkage map of SSC17.

Adenosylhomocysteinase↗

Identification of genomic regions controlling plasma FSH concentrations in Meishan-White Composite boars.

The Chinese Meishan (ME) breed of pig is unique for many reproductive traits. Compared with Western breeds of swine, ME females reach puberty earlier, ovulate more ova per estrus, and have greater uterine capacity, while intact males (boars) have smaller testes and extremely elevated plasma levels of pituitary-derived glycoprotein hormones. In an effort to identify the genetic mechanisms controlling the elevated plasma levels of pituitary-derived glycoprotein hormones [in particular, follicle-stimulating hormone (FSH)] and to determine whether some of these genetic factors are also responsible for differences in other phenotypes, we scanned the entire genome for regions that affected plasma FSH in boars from a Meishan-White Composite (equal contributions of Chester White, Landrace, Large White, and Yorkshire) resource population. Initially, the entire genome of 121 boars was scanned for regions that potentially influenced plasma FSH. The most significant genomic regions were further studied in a total of 436 boars. Three genomic regions located on chromosomes 3, 10, and X apparently possess genes that significantly affect FSH level, and one region provided suggestive evidence for the presence of FSH-controlling genes located on chromosome 8. The region on the X chromosome also affected testes size. Similar genomic regions to those identified on chromosomes 3, 8, and 10 in this study have been identified to affect ovulation rate in female litter mates, supporting the hypothesis that plasma FSH in pubertal boars and ovulation rate in females is controlled by a similar set of genes.

Animals↗