| First Author | Le Tissier PR | Year | 1993 |
| Journal | Mouse Genome | Volume | 91 |
| Issue | 4 | Pages | 879-81 |
| Mgi Jnum | J:16287 | Mgi Id | MGI:64371 |
| Citation | Le Tissier PR, et al. (1993) Mapping studies on distal Chromosome 2. Mouse Genome 91(4):879-81 |
| abstractText | Full text of Mouse Genome contribution: MAPPING STUDIES ON DISTAL CHROMOSOME 2. P.R. Le Tissier l, 2, 3; J.L. Guenet 4, C.J. Skidmore 2 and J. Peters l. 1 MRC Radiobiology Unit, Chilton, Didcot, Oxon, OX11 ORD, UK. 2 Department of Biochemistry & Physiology, University of Reading, PO Box 228, Whiteknights, Reading, RG6 2AJ, UK. 3 Current address: Department of Animal Science, University of Sydney, NSW 2006, Australia. 4 Unite de Genetique des Mammiferes, Institut Pasteur, 25, rue du Docteur Roux, 75724 Paris-Cedex 15, France. Introduction Distal Chromosome (Chr) 2 is one of ten regions of the mouse genome subject to parental imprinting, the phenomenon whereby paternally and maternally derived genes are differentially expressed during development (1). Detailed genetic maps of these regions are useful so that only those genes known with certainty to map to the region are tested for imprinting effects. The region on distal Chr 2 is defined proximally by the T(2;8)2Wa breakpoint in H3 and distally by the T(2;16)28H breakpoint in H4 (2, 3). Current gene markers delimiting the region are adenosine deaminase, Ada, which is in H3 and from linkage analysis maps close to the proximal boundary, and ragged, Ra, which lies close to the distal boundary (4). Wasted, wst, is tightly linked to Ra, for these two loci are estimated to be 0.93 +/- 1.3cM apart (5). Non-agouti, a, is known to map proximal to the region in H1 and is reported to be about 4cM proximal to Ada (6, 7, 8). We report further results of recombinational analysis of genes close to the imprinting region. Materials and Methods All stocks and inbred strains of mice used were from colonies maintained at the MRC Radiobiology Unit, Chilton. DNA samples from the 129XB6 recombinant inbred strain set came from the Institut Pasteur, Paris. ADA allozymes were scored in haemolysates using previously described methods (9). For RFLV scoring of Ada, a full length mouse cDNA clone, pADA5-29 which was a gift of R.E. Kellems, Baylor College, Texas was used (10). Restriction endonuclease digestions, Southern blot analysis and autoradiography were carried out as described previously (11). Results Linkage analysis of wst During earlier experiments (12) a recombinant chromosome, Adab+wstRa, arose and was maintained by crossing to the ADA-B stock, (Adab++/Adab++), to obtain mice of genotype Adab+Ra/Adab++. Preparative matings were then made between Adab+Ra/Adab++ and Adaa+Ra/Adaa wst+; progeny of genotype Adab+Ra/Adaa wst+ were selected, outcrossed to Adaa wst+/Adaa++ and homozygous wst/wst offspring of the outcross were classified for Ada and Ra. The results are shown in Table 1. Of 110 mice classified, no recombinants were found between wst and Ra and thus from the 95% confidence limits wst and Ra are 0-3.24cM apart; when combined with earlier data in which no recombinants were found in 45 mice (12) the loci are within 2.3cM of each other. These estimates are in broad agreement with previous results (5) in which a single recombinant in 461 progeny of a mixed cross, + Ra/ wst + x + +/wst +, was found, putting the distance between the loci at 0.93 +/- 1.3cM. Altogether 26 recombinants occurred between Ada and wst giving a combined recombination frequency of 23.6 +/- 4.0%; no difference in recombination frequency was found between males and females. This figure is somewhat higher than a previous estimate, based on a smaller sample size, of 13.3 +/- 5.1% (12). When both sets of data are combined there have been 32 recombinants in 155 mice giving an RF of 20.6 +/- 3.3%. Table 1: Linkage analysis of Ada and wst. Only wst/wst offspring were informative and are shown here. Parental mating Adab+ Ra/Adaa wst + x Adaa wst +/Adaa+ + Progeny Class: Non-recombinant; Progeny Genotype: Ada: aa; wst: wst wst; Ra: ++; Offspring from Heterozygous: males: 50, females: 34; Combined Number: 84. Progeny Class: Single recombinant, Ada-wst; Progeny Genotype: Ada: ba; wst: wst wst; Ra: ++; Offspring from Heterozygous: males: 15, females: 11; Combined Number: 26. Progeny Class: Single recombinant, wst-Ra; Progeny Genotype: Ada: aa; wst: wst wst; Ra: Ra +; Offsrping from Heterozygous: males: 0, females: 0; Combined Number: 0. Total: 110 Recombination frequencies +/- S.E. (%) Ada-wst: female: 23.1 +/- 5.2; male: 24.4 +/- 6.4; combined: 23.6 +/- 4.0. Linkage analysis of a, Ada and Ra Preparative matings were made between females of genotype + Adab Ra/+ Adab + and C57BL/6J (a Adaa +/a Adaa +). Male offspring of genotype Adab Ra/a Adaa + were selected and backcrossed to female C57BL/6J, and 219 progeny were classified for a, Ra and Ada. The results are shown in Table 2. The gene order, as judged by the rarity of double recombinants, is a Ada Ra for with this order there are none. The RFs between a and Ada of 3.7 +/-1.3% and between Ada and Ra of 17.4 +/- 2.6% are in accord with previously reported values of 4.0 +/- 1.6% (8) and 13.8 +/- 2.7% (12) respectively; the RF of 21.0 +/- 2.8% found in these studies between the anchor loci a and Ra is in agreement with the estimate of 23.7 +/- 1.0% in males (13). Table 2: Linkage analysis of a, Ada and Ra Parental mating a Adaa +/a Adaa + female x + Adab Ra/a Adaa + male. Progeny Class: Non-recombinant: Progeny Genotype: a: + a; Ada: b a; Ra: Ra +; Number: 94; a: a a; Ada: b a; Ra: + +; Number: 79. Progeny Class: Single recombinant a-Ada: Progeny Genotype: a: + a; Ada: a a; Ra: + +; Number: 4; a: a a; Ada: b a; Ra: Ra +; Number: 4. Progeny Class: Single recombinant Ada-Ra: Progeny Genotype: a: + a; Ada: b a; Ra: + +; Number: 16; Progeny Genotype: a: a a; Ada: a a; Ra: Ra +; Number: 22. Total: 219 Recombination frequencies +/- SE (%): a-Ada 3.7 +/- 1.3; Ada - Ra 17.4 +/- 2.6; a Ð Ra: 21.0 +/- 2.8. RI Strain Analysis Genomic DNA digested with PstI and probed with pADA5-29 after Southern blotting showed a 5.6 kb restriction fragment in IS/Cam, 129Sv-SlJ-CP/e, 101/H and ADA-B and a 4.4 kb fragment in C3H/HeH, C57BL/6J, DBA/2Ola/e. The RFLV was scored in 13 129XB RI strains derived from the cross of l29/Sv and C57BL/6JPas. One recombinant was found with non-agouti, a, (Table 3) giving a recombination frequency of 2.17% with upper and lower 95% confidence limits of 0.05 and 19.60% (14). Table 3: Distribution patterns of alleles of Ada and a in the 129XB6 RI strains Strain: C; Locus: a: +; Ada: 129; Strain: D; Locus: a: a; Ada: B; Strain: F; Locus: a: a; Ada: B; Strain: G; Locus: a: +; Ada: 129; Strain: H; Locus a: +; Ada: 129; Strain: I; Locus: a: a; Ada: B; Strain: K; Locus: a: a; Ada: B; Strain: N; Locus: a: +; Ada: 129; Strain: P; Locus: a; Ada: B; Strain: Q; Locus: a: +; X Ada: B; Strain: R; Locus: a: +; Ada: 129; Strain: S; Locus: a: a; Ada: B; Strain: T; Locus: a; Ada: B. Discussion Two studies, the present one and (8), indicate that Ada is about 4cM distal to a, and combining the data from the current work and (12) Ra is 15.9 +/- 1.9cM distal to Ada. These distances accord well with those shown on consensus maps. At the moment Ada and Ra appear to be the closest genes to the boundaries of the region that is subject to imprinting and thus a broad estimate of the genetic length of the region is 16 cM. References 1. Cattanach, B.M. (personal communication). 2. Cattanach, B.M. & Kirk, M. (1985). Nature, 315: 496-498. 3. Cattanach, B.M. et al. (1992). Mouse Genome, 90: 82. 4. Peters, J. et al. (in preparation). 5. Sweet, H.0.(1984). Mouse News Letter, 71: 31. 6. de Boer, P. & van Gijsen, M. (1974). Can. J. Genet. Cytol., 16: 783-788. 7. Washburn, L. & Richer, E.M. (1977). Mouse News Letter, 57: 23. 8. Siracusa, L.D. et al. (1989). Genetics, 122: 669-679. 9. Abbott, C.M. et al. (1986). Proc. Nat. Acad. Sci., USA, 83: 693-695 10. Yeung, C.-Y. et al. (1985). J. Biol. Chem., 260: 10299-10307. 11. Le Tissier, P.R. (1990). Ph.D. thesis, University of Reading. 12. Abbott, C.M. et al. (1991). Biochem. Genet., 29: 537-544. 13. Davisson et al. (1989). In Genetic Variants and Strains of the Laboratory Mouse (eds. M.F. Lyon and A.G. Searle), pp 432-505. Oxford: Oxford University Press. 14. Silver, J. (1985). J. Hered., 76: 436-440. |
