Abstract
Swine Chromosome (Chr) 6-enriched libraries, generated with size-fractionated DNA isolated from chromosomes sorted by flow cytometry, have been used to develop new Chr 6 microsatellite markers. Chromosome isolation procedures were established to reproducibly prepare high quality chromosomes from phytohemagglutinin (PHA)-stimulated swine peripheral blood lymphocytes and to sort individual chromosomes after staining with Hoechst 33258 and chromomycin A3. Chromosome purity was verified by specific staining of swine Chr 6 with fluorescence in situ hybridization (FISH) by use of painting probes generated by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR) amplification of as few as 300 sorted Chr 6. For library construction, DNA was extracted from flow-sorted pools representing Chr 6, amplified, size selected for fragments from 300 to 700 bp, and ligated into pBluescript SK II+ or Lambda ZAP Express. The libraries were then screened with a radiolabeled poly-(dCA) DNA probe. Of 107 (CA)n repeat-containing clones verified by sequencing, 21 were polymorphic and used to genotype the University of Illinois swine reference families. Linkage analysis was then performed with CRIMAP 2.4 (LOD > 3.0), and the results showed that 15 of the microsatellites mapped to swine Chr 6. At least three of these new markers map to locations where there were gaps in the consensus Chr 6 map. Another four markers, because of their PIC values, should provide more informative markers in other areas of the map. Most of the new markers can also be used for automated genotyping with fluorescent labeling. This set of 15 new Chr 6 markers will, therefore, be useful in helping to define QTL associated with swine Chr 6.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ambady S, Mendiola JR, Louis CF, Jansen M, Buoen L, Schook LB, Lunney JK, Grimm DR, Ponce de Leon FA (1997). Development and use of a microdissected swine chromosome 6 DNA library. Cytogenetics & Cell Genetics (in press)
Andersson L, Archibald AL, Gellin J, Schook LB (1993). 1st pig gene mapping workshop (PGM1), 7 August 1992, Interlaken, Switzerland. Anim Genet 24, 205–216
Andersson L, Haley CS, Ellegren H, Knott SA, Johansson M, Andersson K, Andersson-Eklund L, Edfors-Lilja I, Fredholm M, Hansson I, Hakansson J, Lundstrom K (1994). Genetic mapping of quantitative trait loci for growth and fatness in pigs. Science 263, 1771–1774
Archibald AL, Haley CS, Brown JF, Couperwhite S, McQueen HA, Nicholson D, Coppieters W, Van de Weghe A, Stratil A, Wintero A-K, Fredholm M, Larsen NJ, Nielsen VH, Milan D, Woloszyn N, Robic A, Dalens M, Riquet J, Gellin J, Caritez J-C, Burgaud G, Ollivier L, Bidanel J-P, Vaiman M, Renard C, Geldermann H, Davoli R, Ruyter D, Verstege EJM, Groenen MAM, Davies W, Hoyheim B, Keiserud A, Andersson L, Ellegren H, Johansson M, Marklund L, Miller JR, Anderson Dear DV, Signer E, Jeffreys AJ, Moran C, Le Tissier P, Muladno Rothschild MF, Tuggle CK, Vaske D, Helm J, Liu H-C, Rahman A, Yu TP, Larson RG, Schmitz CB (1995). The PiGMaP Consortium linkage map of the pig (Sus scrofa). Mamm Genome, 6, 157–175
Archibald AL, Haley CS, Brown JF, McQueen HA, Lee GJ, Nicholson D, Coppieters W, Van de Weghe A, Wintero A-K, Fredholm M, Nielsen VH, Milan D, Robic A, Riquet J, Gellin J, Davoli R, Groenen MAM, Davies W, Hoyheim B, Andersson L, Signer E, Moran C, Yasue H, Rothschild MF, Tuggle CK, Clutter AC, Pomp D, Grimm DR, Lunney JK, Kirkpatrick B (1997). The PiGMaP Consortium linkage map of the pig (Sus scrofa) — second release. Mammalian Genome (submitted).
Cram LS, Campbell M, Fawcett JJ, Deaven LJ (1990). Polyamine buffer for bivariate human flow cytogenetic analysis and sorting. Methods Cell Biol 33, 377–382
Davies W, Harbitz I, Fries R, Stranzinger G, Hauge JG (1988). Porcine malignant hyperthermia carrier detection and chromosomal assignment using a linked probe. Anim Genet 19, 203–212
Davies W, Hoyheim B, Chaput B, Archibald AL, Frelat G (1994). Characterization of microsatellites from flow-sorted porcine chromosome 13. Mamm Genome 5, 707–711
Dear DVA, Miller JR (1994). Isolation of dinucleotide repeats from a pig chromosome 1-specific DNA library. Mamm Genome 5, 649–651
Echard G, Yerle M, Milan D, Gellin J, Archibald AL (1992). The gene map of the pig (Sus scrofa domestica L.) 2N = 38. In Genetic Maps, Book 4: Nonhuman Vertebrates, 6th edn. (ed. by S.J. O’Brien). (Cold Spring Harbor, N.Y.: Cold Spring Harbor Laboratory
Ellegren H, Basu T (1995). Filling the gaps in the porcine linkage map: isolation of microsatellites from chromosome 18 using flow sorting and SINE-PCR. Cytogenet Cell Genet 71, 370–373
Ellegren H, Johansson M, Chowdhary BP, Marklund S, Ruyter D, Marklund L, Brauner-Nielsen P, Edfors-Lilja I, Gustavsson I, Juneja RK, Andersson L (1993). Assignment of 20 microsatellite markers to the porcine linkage map. Genomics 16, 431–439
Ellegren H, Chowdhary B, Johansson M, Andersson L (1994). Integrating the porcine physical and linkage map using cosmid-derived markers. Anim Genet 25, 155–164
Fredholm M, Wintero AK, Christensen K, Kristensen B, Brauner-Nielsen P, Davies W, Archibald AL (1993). Characterization of 24 porcine (dA-dC)n-(dT-dG)n microsatellites: genotyping of unrelated animals from four breeds and linkage studies. Mamm Genome 4, 187–192
Frengen E, Thomsen PD, Schmitz A, Frelat G, Davies W (1994). Isolation of region-specific probes from pig chromosome 6 by coincidence cloning. Mamm Genome 5, 497–502
Fuji J, Otsu K, Zorzato F, deLeon S, Khanna VK, Weiler JE, O’Brien PJ, MacLennan DH (1991). Identification of a mutation in porcine ryano-dine receptor associated with malignant hyperthermia. Science 253, 448–451
Green P, Falls K, Crooks S (1990). Documentation for CRI-MAP, Version 2.4. (St. Louis, Mo.: Washington University School of Medicine)
Harbitz I, Chowdhary B, Thomsen PD, Davies W, Kaufmann U, Kran S, Gustavsson I, Christensen K, Hauge JG (1990). Assignment of the porcine calcium release channel gene, a candidate for the malignant hyperthermia locus, to the 6p11>q21 segment of chromosome 6. Genomics 8, 243–248
Hughes IP, Moran C, Nicholas FW (1992). PCR genotyping of the ryano-dine receptor gene for a putative causal mutation for malignant hyperthermia in Australian pigs. J Anim Breed Genet 109, 465–476
Jensen P, Barton-Gade P (1985). Performance and carcass characteristics of pigs with known genotypes for halothane susceptibility. Eur Assn Anim Prod Pub 33, 80–87
Langford CF, Telenius H, Miller NGA, Thomsen PD, Tucker EM (1993). Preparation of chromosome-specific paints and complete assignment of chromosomes in the pig flow karyotype. Anim Genet 24, 261–267
Lichter P, Cremer T (1992). Chromosome analysis by non-isotopic in situ hybridization. In Human Cytogenetics, Vol. I, Constitutional Analysis, (DE Rooney, BH Czepulkowski, eds.). Oxford, New York, Tokyo:, I.R.L., pp. 157–192
Lincoln SE, Daly MJ, Lander ES (1991). PRIMER: A Computer Program for Automatically Selecting PCR Primers. (Cambridge, Mass.: MIT Center for Genome Research and Whitehead Institute for Biomedical Research)
Mariani P, Lundstrom K, Gustafsson U, Enfalt A-C, Juneja RK, Andersson L (1996). A major locus (RN) affecting muscle glycogen content is located on pig chromosome 15. Mamm Genome 7, 52–54
Milan D, Woloszyn N, Yerle M, Le Roy P, Bonnet M, Riquet J, Lahbib-Mansais Y, Caritez J-C, Robic A, Sellier P, Elsen J-M, Gellin J (1996). Accurate mapping of the “acid meat” RN gene on genetic and physical maps of pig chromosome 15. Mamm Genome 7, 47–51
Miller JR, Dixon SC, Miller NGA, Tucker EM, Hindkjaer J, Thomsen PD (1992). A chromosome 1-specific DNA library from the domestic pig (Sus scrofa domestica). Cytogenet Cell Genet 61, 128–131
Otsu K, Khanna VK, Archibald AL, MacLennan DH (1991). Cosegregation of porcine malignant hyperthermia and a probable causal mutation in the skeletal muscle ryanodine receptor gene in backcross families. Genomics 11, 744–750
Paszek AA, Schook LB, Louis CF, Mickelson JR, Flickenger GH, Murtaugh MA, Mendiola J, Beattie CW, Rohrer GA, Alexander LJ, Andersson L, Ellegren H, Johansson M, Mariani P, Marklund L, Hoyheim B, Davies W, Fredholm M, Archibald AL, Haley CS (1995). First International workshop on porcine chromosome 6. Anim Genet 26:377–401.
Rettenberger G, Fries R, Enge W, Scheit KH, Dolf G, Hameister H (1994). Establishment of a partially informative porcine somatic cell hybrid panel and assignment of the loci for transition protein 2 (TNP2) and protamine 1 (PRM1) to chromosome 3 and polyubiquitin (UBC) to chromosome 14. Genomics 21, 558–566
Riquet J, Milan D, Woloszyn N, Schmitz A, Pitel F, Frelat G, Gellin J (1995). A linkage map with microsatellites isolated from swine flow-sorted chromosome 11. Mamm Genome 9, 623–628
Riquet J, Schmitz A, Cruzel J, Mulsant P, Milan D, Gellin J (1996). Assignment of markers by using polymerase chain reaction on pools of swine flow-sorted chromosomes. Anim Genet 27, 133–136
Robic A, Riquet J, Yerle M, Milan D, Lahbib-Mansais Y, Dubut-Fontana C, Gellin J (1996). Porcine linkage and cytogenetic maps integrated by regional mapping of 100 microsatellites on somatic cell hybrid panel. Mamm Genome 7, 438–445
Rohrer GA, Alexander LJ, Keele JW, Smith TP, Beattie CW (1994). A microsatellite linkage map of the porcine genome. Genetics 136, 231–245
Rohrer GA, Alexander LJ, Hu Z, Smith TPL, Keele JW, Beattie CW (1996). A comprehensive map of the porcine genome. Genome Res 6, 371–391
Rothschild MF, Jacobson C, Vaske DA, Tuggle C, Wang L, Short T, Eckardt, G, Sasaki S, Vincent A, McLaren DG, Southwood O, van der Steen H, Mileham A, Plastow G (1996). The estrogen receptor locus is associated with a major gene influencing litter size in pigs. Proc Natl Acad Sci USA 93, 201–205
Schmitz A, Chaput B, Fouchet P, Guilly MN, Frelat G, Vaiman M (1992). Swine chromosomal DNA quantification by bivariate flow karyotyping and karyotype interpretation. Cytometry 13, 703–710
Simpson SP, Webb AJ (1989). Growth and carcass performance of British Landrace pigs heterozygous at the halothane locus. Anim Prod 49, 503
Stallings RL, Ford AF, Nelson D, Torney DC, Hildebrand CE, Moyzis RK (1991). Evolution and distribution of (GT)n repetitive sequences in mammalian genomes. Genomics 10, 807–815
Telenius H, Carter NP, Bebb CE, Nordenskjold M, Ponder BAJ, Tunnac-liffe A (1992). Degenerate oligonucleotide-primed PCR: general amplification of target DNA by a single degenerate primer. Genomics 13, 718–725
Wintero AK, Fredholm M, Thomsen PD (1992). Variable (dG-dT)n-(dC-dA)n sequences in the porcine genome. Genomics 12, 281–288
Zhang TQ, Buoen LC, Weber AF, Christianson B, Morrison R, Marsh W, Ruth GR (1992). A chromosomal reciprocal translocation (lq+; 14q-) in a boar siring reduced litter sizes. Theriogenology 38, 799–806
Zhang W, Kuhlers DL, Rempel WE (1992). Halothane gene and swine performance. J Anim Sci 70, 1307–1313
Zijlstra C, Bosma AA, de Haan NA, Mellink C (1996). Construction of a cytogenetically characterized porcine somatic cell hybrid panel and its use as a mapping tool. Mamm Genome 7, 280–284
Author information
Authors and Affiliations
Additional information
The nucleotide sequence data reported in this paper have been submitted to GenBank and assigned the accession numbers U24283, U24284, U24285, U24286, U62944, U62945, U62946, U62947, U62948, U62949, U62950, U62951, U62952, U62953, U62954, U62955, U62956, U62957, U62958, U62959, and U66700.
The U.S. Government has a right to obtain a non-exclusive royalty-free license to any copyright covering this paper.
Rights and permissions
About this article
Cite this article
Grimm, D.R., Goldman, T., Holley-Shanks, R. et al. Mapping of microsatellite markers developed from a flow-sorted swine chromosome 6 library. Mammalian Genome 8, 193–199 (1997). https://doi.org/10.1007/s003359900388
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s003359900388