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Additional Linkage Relationships Within the Z Chromosome of the Chicken1,2
Additional Linkage Relationships Within the Z Chromosome of the Chicken1'2 J.J.BITGOOD Department of Poultry Science, University of Wisconsin-Madison, Madison, Wisconsin 53706 (Received for publication November 30, 1984)
1985 Poultry Science 64:2234-2238 INTRODUCTION Classical Mendelian linkage studies have defined t h e linkage relationships of loci located o n t h e Z c h r o m o s o m e in t h e chicken, as recently summarized by Somes ( 1 9 8 4 ) . Beginning with Z a r t m a n ' s r e p o r t ( 1 9 7 3 ) on t h e i n d u c t i o n of t w o Z-linked c h r o m o s o m e translocations by X-irradiation of semen and assignment of t h e pea c o m b locus t o c h r o m o s o m e 1, additional techniques from t h e field of cytogenetics have b e c o m e available for chicken linkage studies. Telloni et al. ( 1 9 7 6 ) were t h e first t o d e t e r m i n e t h e m a p distance between a Z-linked locus and a c h r o m o s o m e translocation break p o i n t in t h e chicken. In a detailed analysis of a t ( Z ; m i c r o ) interchange, t h e y found t h a t t h e Z-linked rate of feathering locus (K) was 23 m a p units from t h e break point. T h e y also reported t h a t t h e Z-linked barring locus (B) was n o t on t h e arm of t h e c h r o m o s o m e t h a t contained t h e break p o i n t .
1 Part of a contributing project to the North Central Breeding Project, NC-168. Supported in part by the College of Agricultural and Life Sciences, University of Wisconsin, Madison and by The Graduate School, University of Wisconsin, Madison, Project No. 150364. 2 Portions of this project were presented at the 1984 annual meeting of the Poultry Science Association, University of Guelph, Guelph, Ontario.
Bitgood et al. ( 1 9 8 0 ) utilized t w o Z-linked translocations, t h e MN t ( Z ; l ) , induced b y Wang et al. ( 1 9 8 2 ) , and t h e NM 7 0 9 2 t ( Z ; l ) , induced b y Z a r t m a n ( 1 9 7 3 ) , t o investigate further t h e linkage relationship between B and Z-linked silver (S). T h a t study also provided evidence t h a t B and S were on opposite arms of the Z c h r o m o s o m e . Tentative arm assignments were m a d e ; however, sufficient i n f o r m a t i o n was n o t available t o confirm these designations. This paper presents additional linkage information for several c o m b i n a t i o n s of four Z-linked loci and t h r e e Z-linked c h r o m o s o m e rearrangements and reviews several o t h e r studies t h a t n o w appear t o p e r m i t assignment of these markers t o specific a r m s of t h e Z c h r o m o s o m e in the chicken. MATERIALS AND METHODS T h r e e Z-linked c h r o m o s o m e translocations were utilized in these tests. T h e MN t ( Z ; l ) has been previously described (Wang et al., 1 9 8 2 ; Bitgood, 1 9 8 0 ) . T h e NM 7 6 5 9 t ( Z ; l ) was first described b y Z a r t m a n ( 1 9 7 3 ) and later by Kaelbling and F e c h h e i m e r (1983b), who designated it t h e t(NM 1). T h e MN t(Z;3) was described b y Wang et al. ( 1 9 8 2 ) . F o u r test matings were c o n d u c t e d : Barring, Silver, Recessive White Skin. Males with standard c h r o m o s o m e s and h o m o z y g o u s for Z-linked barring (B), silver (S), and yellow
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ABSTRACT Genetic linkage tests were conducted using four Z-linked loci and three Z-linked chromosome rearrangements (interchanges). Z-linked recessive white skin (y) segregated independently from the Z-linked barring (B) and silver (S) loci. A map distance of 18.2 ± 3.2 was found between y and the MN t(Z;l) interchange. A previous report had shown that B was located 22 map units from this interchange, therefore, because B and y were found not to show measurable recombination, the linear order is B-interchange-y. The MN t(Z-,3) interchange was tested against S, and no recombinants were recovered in 217 backcross progeny, indicating very close genetic linkage. The NM 7659 t(Z;l) interchange was tested against the dermal melanin locus (id+) and independent assortment was noted. This supports other work that has shown that the S and id+ loci lie on opposite arms of the Z chromosome. Evidence is reviewed that supports the concept that S and the NM 7659 and MN t(Z;3) interchange break points are located on the short arm, while B, id+, y, and the MN t(Z;l) interchange break point are located on the long arm of the Z chromosome. (Key words: chicken, Z-linkage, gene mapping, translocation (interchange) mapping)
Z CHROMOSOME LINKAGES
information about the segregation of silver and gold. NM 7659 t(Z;l), Dermal Melanin. Araucana males with standard chromosomes and homozygous for Z-linked dermal melanin (id+) were mated to females hemizygous for dermal melanin inhibitor (Id) and the NM 7659 t(Z;l). The Fj males were backcrossed to Single Comb White Leghorn females with standard chromosomes and hemizygous for Id. The backcross progeny were reared to 6 weeks, at which time feather pulp samples for chromosome analysis were obtained, and phenotypic descriptions recorded. Only female progeny could be scored for segregation at the Id locus. These females were also used in another study, and when they were placed in floor laying pens at 18 weeks of age, the shank color of each bird was verified. With the exception of the day-old black chicks in the MN t(Z;3) mating discussed above, phenotypic descriptions were obtained after feather pulp analysis to prevent any unconscious bias in case of a questionable analysis. Samples were repeated in any cases of questionable analysis. Segregation data were analyzed following procedures outlined by Green (1963). RESULTS The segregations and analysis of the B-S-Y+l b -s+-y X b+-s+-y/-w backcross matings are presented in Table 1. The numbers differ between the B-y and S-y analysis because silver and gold could not be accurately identified in barred birds in a black background (Campo and Orozco, 1980). There were no significant +
TABLE 1. Tests for the linkage of Z-linked recessive white skin (y) with Z-linked barring (B) and silver (S) n
Gamete type
92 94 70 92 348 Chi-square X2B=
s-v+o s+-y<> S+-Y+
s-y
n 63 65 46 74 248
1-15
x2s=
2.73
X^,=
1-66
X^=
3.63
X2L=
1-66
x^=
Parental gametic types.
.26
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skin (Y+) were mated to females with standard chromosomes and hemizygous for nonbarring (b+), gold (s + ), and Z-linked recessive white skin (y) (McGibbon, 1981). The F! males were then backcrossed to b+, s+, y females with Standard chromosomes. All backcross progeny were reared to 6 weeks of age, at which time phenotypic descriptions were obtained. MN t(Z;l), Recessive White Skin. Males homozygous for yellow skin (Y+/Y+) and the MN t(Z;l) were mated to females with standard chromosomes and hemizygous for y. The Fj males were backcrossed to y females with standard chromosomes. The backcross progeny were reared to 6 weeks of age, at which time feather pulp samples for chromosome analyses were obtained (Shoffner et al., 1967). Phenotypic descriptions were recorded after chromosome complements were determined. MN t(Z;3), Barring, Silver. Males with standard chromosomes and homozygous for B and S were mated to females hemizygous for b+, s + , and the MN t(Z;3). The F t males were backcrossed to females with standard chromosomes and hemizygous for b+ and s + . Campo and Orozco (1980) had reported difficulty in accurately identifying the silver and gold feather patterns in black-barred birds. Because of this difficulty, it was originally decided for this study not to attempt to analyze the segregation of S and x + in black chicks recovered from this mating. All black chicks from the first two hatches were scored for B or b+ at hatch time; feather pulp samples for chromosome analysis were obtained and these chicks were then discarded. A modification of the feather pulp procedure described by Shoffner et al. (1967) was used on the day-old chicks. An injection of .06 ml of a .05% (w/v) colchicine solution was injected subcutaneously in the back of the neck (as in the usual vaccination procedure for Marek's disease), instead of an interperitoneal injection. All other steps in the procedure were unchanged. The remaining chicks were reared to 6 weeks of age for chromosome analysis and phenotypic descriptions. Prior to the third hatch from this mating, phenotypic descriptions of progeny from the B-S-y test mating described were obtained. It was noted that the black nonbarred chicks were apparently birchen black, as silver or gold lacing was seen in the hackles of these juveniles. Therefore, all chicks from the remaining hatches of the MN t(Z;3 )-B-S test mating were reared to 6 weeks for additional
2235
BITGOOD
2236
TABLE 2. Linkage between the MN t(Z;l) and Z-linked recessive white skin (y) Gamete type
n
Chi-square
X—Y + < >
83
X2T=
N-y°
47
X^ = 7.70*
N-K+
14
X? =64.16*** Contingency x [ = 60.25***
15 159
T-y
8.61***
Map distance = 18.2 ± 3.2 Parental gametic type; T = rearrangement carrier, N = normal karyotype. *P<.01. ***P<.005.
TABLE 3. Tests for linkage of the MN t(Z;3) with Z-linked barring (B) and silver (S) Gamete type T-b«>
N-rf> N-b+ T-B
n
Gamete type
52 75 60 67 254
T-s-'O N-SO N-s+ T-S
4= 4=
3.54
99 118 0 0 217
.00
*P<.005.
1.66
4= 4=
DISCUSSION Data from a series of reports now make it possible to assign several Z-linked loci to specific arms of the Z chromosome. Kaelbling and Fechheimer (1983a) reported on comparative studies of synaptonemal complexes (SC) formed in normal chicken males, and also (SC) formed in normal chicken males, and also SC formed by males heterozygous for the NM Zartman in 1973 (Kaelbling and Fechheimer, 1983b). They found that this translocation break point is on the arm of the Z chromosome that contains the K and dwarfing (dw) loci, and that B and the terminal C-band are on the arm that is not involved in this translocation. The data presented in Table 4 supports this, as the NM 7659 segregates independently from id+, and id+ has been mapped ca. 10 map units from B (Punnett, 1940). Telloni et al. (1976) studied the t(OH 10), a translocation between the Z and a microchromosome, and found that the K and dw loci were located on the arm that contained the break point. The B locus showed no linkage with the break point. Blazak and Fechheimer (1980) conducted a C-band analysis of this same translocation and reported that the terminal C-band (G-band light staining region) on the Z chromosome was located on the intact arm. Kaelbling and Fechheimer (1983b) obtained measurements of the synaptonemal complex
n
Chi-square 1.01
recovered, showing that extremely tight genetic linkage exists between these markers. Data for the NM 7659 X id+ (Z-linked dermal melanin) mating are presented in Table 4. There was no significant linkage relationship shown between these markers.
TABLE 4. Test for linkage of the NhI 7659 with Z-linked dermal melanin
t(Z;l)
Gamete type
n
Chi-square
T-IdO
58
XT=1.67
U-id+0
54
4 r -60 xL-l-07
1.66
N-Id
56
217.0***
T-id+
72 240
Parental gametic type; T = rearrangement carrier, N = normal karyotype.
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linkage relationships found between y and either B or S. The data for the MN t(Z;l) X y backcross mating are shown in Table 2. A significant linkage of 18.2 ± 3.2 map units was calculated between these markers. The data for the MN t(Z;3) X B-S backcross mating are presented in Table 3. As described in the Materials and Methods section, the numbers differ between the B and the S analyses, because black chicks from the first two hatches were not reared to determine segregation at the S locus. Independent assortment was noted between the break point and B. No recombinants between the break point and S were
Z CHROMOSOME LINKAGES
arm opposite the arm containing the terminal G-band light staining region, again showing that the S-K-dw region is on the p arm of the Z chromosome. This MN (Z;3)-S intimate relationship is the third interchange/morphological trait relationship of this type that has been found in the chicken. The first reported was the NM 7092 translocation (Zartman, 1973), which showed complete linkage to a protoporphyrin mutant (pr) (Shoffner et al, 1982). The second was complete linkage of a pericentric inversion within chromosome 2 with a limb development mutation that was "locked into" the inverted segment (Langhorst and Fechheimer, 1983). Specific regions of three chromosomes have now been defined by both cytological and morphological markers. The definition of the By region of the q arm of the Z chromosome will be useful for further linkage studies. These two easily identifiable markers are now mapped in relation to a common, almost central marker (the MN t(Z;l)), and have extended the sweep distance for genetic testing of this region of the q arm to greater than 50 map units. The id+ locus, known to be linked to B by 10 map units, will be tested against the MN t(Z;l) to attempt to determine its location within this chromosome region. Also, F t males have been obtained that are genotypically id+-B-y/Id-b+-Y+ in order to attempt to further refine linkage relationships between Id, B, and Y+ in this region. Results from this mating should also disclose whether or not y will interact with id+ in a manner similar to the interaction of id+ and autosomal dominant white skin (W+), which results in blue or slaty blue/black shanks (Hutt, 1949). While independence was noted between B and y, use of the MN t(Z;l) interchange showed that these loci are in fact located on the same arm of the Z chromosome, another indication of the apparently extremely long genetic distances in the larger chicken chromosomes. Punnett (1940) also discussed this in a review of data gathered by himself and several other investigators. At that time, the possibility that the largest chromosome was the sex chromosome was being discussed. It is now known that the Z chromosome is the fifth largest in the chicken, and its genetic length is apparently longer than the average length of any chromosome of the laboratory mouse (Womack, 1979). This reinforces the importance of utilizing
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formed by this t(OH 10) rearrangement and determined that the arm that was involved in the rearrangement was the short arm (Fechheimer, personal communication). The results of these four studies indicate that the G-band light staining region of the Z chromosome marks the long (q) arm of this chromosome. The break point on the Z chromosome in the MN t(Z;l) is in the G-band light staining region (Bitgood, 1980). This break point, therefore, is on the q arm of the Z chromosome. Previous mapping of this same rearrangement against B showed 22 map units between these markers (Bitgood etal, 1980). The linkage of 18 map units shown between this break point and y (Table 2) and the independence found between B and y (Table 1), suggest the linear order of £(22)T(18)y (T representing the translocation break point) on the long arm of the Z chromosome. Females hemizygous for B-y have been mated to homozygous MN t(Z;l) males to obtain F! males that will be used in a three point test cross to verify these findings. It is not yet possible to determine which direction the centromere or the telomere lie from this chromosome segment. Significant deviations from expected numbers of Y+ and y individuals are noted in Table 2. Sixty-two individuals were scored as y and 97 were scored as Y+. In the same table, there is also a deviation from expected in the segregation of the translocation, 50 normals and 84 translocation carriers. These are possibly chance deviations, as other matings using either the translocation or y have segregated normally (unpublished observations). The results from the future test mating involving the MN t(Z;l), B, and y (described above) will be monitored to determine if any unusual segregations might be occurring in this chromosomal region. A contingency chi-square for linkage was calculated using the procedure presented by Green (1963). From the results of the MN t(Z;3) X B, S backcross mating, it is evident that this break point is genetically very closely linked to S. Due to the reduction in crossing over expected in the vicinity of an interchange, it is not known what the physical distance between these markers might be. Further tests of this rearrangement against other loci on the short (p) arm of the chromosome will clarify this relationship. Wang and Shoffner (1974) conducted G-band analysis of this interchange and demonstrated that the break point was on the
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BITGOOD
multiple point test crosses in chicken gene linkage studies in order to define regions for expansion of sweep distances for additional linkage tests. ACKNOWLEDGMENTS
REFERENCES Bitgood, J. J., 1980. Gametogenesis and zygote formation in domestic fowl (Gallus domesticus) with chromosome rearrangements. Ph.D. Thesis, University of Minnesota, St. Paul, MN. Bitgood, J. J., R. N. Shoffner, J. S. Otis, and W. E. Briles, 1980. Mapping of the genes for pea comb, blue egg, barring, silver, and blood groups A, E, H, and P in the domestic fowl. Poultry Sci. 59:1686- 1693. Blazak, W. F., and N. S. Fechheimer, 1980. Gonosome-autosome translocations in fowl: meiotic configurations and chiasma counts from singly and doubly heterozygous cockerals. Can. J. Genet. Cytol. 22:343-351.. Campo, J. L., and F. Orozco, 1980. The action of the sex linked barring gene on Spanish chickens with gold plumage. Ann. Genet. Sel. Anim. 12: 233-239. Green, M. C , 1963. Methods for testing linkage. Pages 56—82 in Methodology in Mammalian Genetics. W. J. Burdette, ed. Holden-Day, Inc., San Francisco, CA. Hutt, F. B., 1949. Genetics of the Fowl. McGraw-Hill Book Company, Inc., New York, NY. Kaelbling, M., and N. S. Fechheimer, 1983a. Synaptonemal complexes and the chromosome complement of domestic fowl, Gallus domesticus. Cytogenet. Cell Genet. 35:87-92.
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Appreciation is expressed to R. N. Shoffner, University of Minnesota and N. S. Fechheimer, The Ohio State University for furnishing the chromosome rearrangement lines used in this study. Appreciation is also expressed to L. Neumann, W. Schumann, E. Eutsler, and B. Rhiner for technical assistance.
Kaelbling, M., and N. S. Fechheimer, 1983b. Synaptonemal complex analysis of chromosome rearrangements in domestic fowl, Gallus domesticus. Cytogenet. Cell Genet. 36:567— 572. Langhorst, L. J., and N. S. Fechheimer, 1983. Correlation between presence of a pericentric inversion of the second chromosome and limb phenotype in the domestic chicken. J. Dairy Sci. 66 (Suppl. 1):246. (Abstr.) McGibbon, W. H., 1981. White skin: a Z-linked recessive mutation in the fowl. J. Hered. 72: 139-140. Punnett, R. C , 1940. Genetic studies in poultry. X. Linkage data for the sex chromosome. J. Genet. 39:335-342. Shoffner, R. N., A. Krishan, G. J. Haiden, R. K. Bammi, and J. S. Otis, 1967. Avian chromosome methodology. Poultry Sci. 56:334—344. Shoffner, R. N., R. Shuman, J. S. Otis, J . J . Bitgood, V. Garwood, and P. Lowe, 1982. The effect of a protoporphyrin mutant on some economic traits of the chicken. Poultry Sci. 61:817-820. Somes, R. G., Jr., 1984. Linked loci of the chicken — Gallus gallus (G. domesticus). Genet. Maps 3:465-473. Telloni, R. V., R. G. Jaap, and N. S. Fechheimer, 1976. Cytogenetic and phenotypic effects of a chromosomal rearrangement involving the Zchromosome and a micro-chromosome in the chicken. Poultry Sci. 55:1886-1896. Wang, N., and R. N. Shoffner, 1974. Trypsin G- and C-banding for interchange analysis and sex identification in the chicken. Chromosoma (Berl.) 47:61-69. Wang, N., R. N. Shoffner, J. S. Otis, and K. M. Cheng, 1982. The induction of chromosomal structural changes in male chickens by the alkylating agents: triethylene melamine and ethyl methanesulfonate. Mutat. Res. 96:53—66. Womack, J. E., 1979. Linkage map: mouse. Pages 38—39 in Inbred and Genetically Defined Strains of Laboratory Animals. Part 1. Mouse and Rat. P. L. Altman and D. D. Katz, ed. Fed. Am. Soc. Exper. Biol., Bethesda, MD. Zartman, D. L., 1973. Location of the pea comb gene. Poultry Sci. 52:1455-1462.
1985 Poultry Science 64:2234-2238 INTRODUCTION Classical Mendelian linkage studies have defined t h e linkage relationships of loci located o n t h e Z c h r o m o s o m e in t h e chicken, as recently summarized by Somes ( 1 9 8 4 ) . Beginning with Z a r t m a n ' s r e p o r t ( 1 9 7 3 ) on t h e i n d u c t i o n of t w o Z-linked c h r o m o s o m e translocations by X-irradiation of semen and assignment of t h e pea c o m b locus t o c h r o m o s o m e 1, additional techniques from t h e field of cytogenetics have b e c o m e available for chicken linkage studies. Telloni et al. ( 1 9 7 6 ) were t h e first t o d e t e r m i n e t h e m a p distance between a Z-linked locus and a c h r o m o s o m e translocation break p o i n t in t h e chicken. In a detailed analysis of a t ( Z ; m i c r o ) interchange, t h e y found t h a t t h e Z-linked rate of feathering locus (K) was 23 m a p units from t h e break point. T h e y also reported t h a t t h e Z-linked barring locus (B) was n o t on t h e arm of t h e c h r o m o s o m e t h a t contained t h e break p o i n t .
1 Part of a contributing project to the North Central Breeding Project, NC-168. Supported in part by the College of Agricultural and Life Sciences, University of Wisconsin, Madison and by The Graduate School, University of Wisconsin, Madison, Project No. 150364. 2 Portions of this project were presented at the 1984 annual meeting of the Poultry Science Association, University of Guelph, Guelph, Ontario.
Bitgood et al. ( 1 9 8 0 ) utilized t w o Z-linked translocations, t h e MN t ( Z ; l ) , induced b y Wang et al. ( 1 9 8 2 ) , and t h e NM 7 0 9 2 t ( Z ; l ) , induced b y Z a r t m a n ( 1 9 7 3 ) , t o investigate further t h e linkage relationship between B and Z-linked silver (S). T h a t study also provided evidence t h a t B and S were on opposite arms of the Z c h r o m o s o m e . Tentative arm assignments were m a d e ; however, sufficient i n f o r m a t i o n was n o t available t o confirm these designations. This paper presents additional linkage information for several c o m b i n a t i o n s of four Z-linked loci and t h r e e Z-linked c h r o m o s o m e rearrangements and reviews several o t h e r studies t h a t n o w appear t o p e r m i t assignment of these markers t o specific a r m s of t h e Z c h r o m o s o m e in the chicken. MATERIALS AND METHODS T h r e e Z-linked c h r o m o s o m e translocations were utilized in these tests. T h e MN t ( Z ; l ) has been previously described (Wang et al., 1 9 8 2 ; Bitgood, 1 9 8 0 ) . T h e NM 7 6 5 9 t ( Z ; l ) was first described b y Z a r t m a n ( 1 9 7 3 ) and later by Kaelbling and F e c h h e i m e r (1983b), who designated it t h e t(NM 1). T h e MN t(Z;3) was described b y Wang et al. ( 1 9 8 2 ) . F o u r test matings were c o n d u c t e d : Barring, Silver, Recessive White Skin. Males with standard c h r o m o s o m e s and h o m o z y g o u s for Z-linked barring (B), silver (S), and yellow
2234
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ABSTRACT Genetic linkage tests were conducted using four Z-linked loci and three Z-linked chromosome rearrangements (interchanges). Z-linked recessive white skin (y) segregated independently from the Z-linked barring (B) and silver (S) loci. A map distance of 18.2 ± 3.2 was found between y and the MN t(Z;l) interchange. A previous report had shown that B was located 22 map units from this interchange, therefore, because B and y were found not to show measurable recombination, the linear order is B-interchange-y. The MN t(Z-,3) interchange was tested against S, and no recombinants were recovered in 217 backcross progeny, indicating very close genetic linkage. The NM 7659 t(Z;l) interchange was tested against the dermal melanin locus (id+) and independent assortment was noted. This supports other work that has shown that the S and id+ loci lie on opposite arms of the Z chromosome. Evidence is reviewed that supports the concept that S and the NM 7659 and MN t(Z;3) interchange break points are located on the short arm, while B, id+, y, and the MN t(Z;l) interchange break point are located on the long arm of the Z chromosome. (Key words: chicken, Z-linkage, gene mapping, translocation (interchange) mapping)
Z CHROMOSOME LINKAGES
information about the segregation of silver and gold. NM 7659 t(Z;l), Dermal Melanin. Araucana males with standard chromosomes and homozygous for Z-linked dermal melanin (id+) were mated to females hemizygous for dermal melanin inhibitor (Id) and the NM 7659 t(Z;l). The Fj males were backcrossed to Single Comb White Leghorn females with standard chromosomes and hemizygous for Id. The backcross progeny were reared to 6 weeks, at which time feather pulp samples for chromosome analysis were obtained, and phenotypic descriptions recorded. Only female progeny could be scored for segregation at the Id locus. These females were also used in another study, and when they were placed in floor laying pens at 18 weeks of age, the shank color of each bird was verified. With the exception of the day-old black chicks in the MN t(Z;3) mating discussed above, phenotypic descriptions were obtained after feather pulp analysis to prevent any unconscious bias in case of a questionable analysis. Samples were repeated in any cases of questionable analysis. Segregation data were analyzed following procedures outlined by Green (1963). RESULTS The segregations and analysis of the B-S-Y+l b -s+-y X b+-s+-y/-w backcross matings are presented in Table 1. The numbers differ between the B-y and S-y analysis because silver and gold could not be accurately identified in barred birds in a black background (Campo and Orozco, 1980). There were no significant +
TABLE 1. Tests for the linkage of Z-linked recessive white skin (y) with Z-linked barring (B) and silver (S) n
Gamete type
92 94 70 92 348 Chi-square X2B=
s-v+o s+-y<> S+-Y+
s-y
n 63 65 46 74 248
1-15
x2s=
2.73
X^,=
1-66
X^=
3.63
X2L=
1-66
x^=
Parental gametic types.
.26
Downloaded from http://ps.oxfordjournals.org/ at University of Georgia on May 25, 2015
skin (Y+) were mated to females with standard chromosomes and hemizygous for nonbarring (b+), gold (s + ), and Z-linked recessive white skin (y) (McGibbon, 1981). The F! males were then backcrossed to b+, s+, y females with Standard chromosomes. All backcross progeny were reared to 6 weeks of age, at which time phenotypic descriptions were obtained. MN t(Z;l), Recessive White Skin. Males homozygous for yellow skin (Y+/Y+) and the MN t(Z;l) were mated to females with standard chromosomes and hemizygous for y. The Fj males were backcrossed to y females with standard chromosomes. The backcross progeny were reared to 6 weeks of age, at which time feather pulp samples for chromosome analyses were obtained (Shoffner et al., 1967). Phenotypic descriptions were recorded after chromosome complements were determined. MN t(Z;3), Barring, Silver. Males with standard chromosomes and homozygous for B and S were mated to females hemizygous for b+, s + , and the MN t(Z;3). The F t males were backcrossed to females with standard chromosomes and hemizygous for b+ and s + . Campo and Orozco (1980) had reported difficulty in accurately identifying the silver and gold feather patterns in black-barred birds. Because of this difficulty, it was originally decided for this study not to attempt to analyze the segregation of S and x + in black chicks recovered from this mating. All black chicks from the first two hatches were scored for B or b+ at hatch time; feather pulp samples for chromosome analysis were obtained and these chicks were then discarded. A modification of the feather pulp procedure described by Shoffner et al. (1967) was used on the day-old chicks. An injection of .06 ml of a .05% (w/v) colchicine solution was injected subcutaneously in the back of the neck (as in the usual vaccination procedure for Marek's disease), instead of an interperitoneal injection. All other steps in the procedure were unchanged. The remaining chicks were reared to 6 weeks of age for chromosome analysis and phenotypic descriptions. Prior to the third hatch from this mating, phenotypic descriptions of progeny from the B-S-y test mating described were obtained. It was noted that the black nonbarred chicks were apparently birchen black, as silver or gold lacing was seen in the hackles of these juveniles. Therefore, all chicks from the remaining hatches of the MN t(Z;3 )-B-S test mating were reared to 6 weeks for additional
2235
BITGOOD
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TABLE 2. Linkage between the MN t(Z;l) and Z-linked recessive white skin (y) Gamete type
n
Chi-square
X—Y + < >
83
X2T=
N-y°
47
X^ = 7.70*
N-K+
14
X? =64.16*** Contingency x [ = 60.25***
15 159
T-y
8.61***
Map distance = 18.2 ± 3.2 Parental gametic type; T = rearrangement carrier, N = normal karyotype. *P<.01. ***P<.005.
TABLE 3. Tests for linkage of the MN t(Z;3) with Z-linked barring (B) and silver (S) Gamete type T-b«>
N-rf> N-b+ T-B
n
Gamete type
52 75 60 67 254
T-s-'O N-SO N-s+ T-S
4= 4=
3.54
99 118 0 0 217
.00
*P<.005.
1.66
4= 4=
DISCUSSION Data from a series of reports now make it possible to assign several Z-linked loci to specific arms of the Z chromosome. Kaelbling and Fechheimer (1983a) reported on comparative studies of synaptonemal complexes (SC) formed in normal chicken males, and also (SC) formed in normal chicken males, and also SC formed by males heterozygous for the NM Zartman in 1973 (Kaelbling and Fechheimer, 1983b). They found that this translocation break point is on the arm of the Z chromosome that contains the K and dwarfing (dw) loci, and that B and the terminal C-band are on the arm that is not involved in this translocation. The data presented in Table 4 supports this, as the NM 7659 segregates independently from id+, and id+ has been mapped ca. 10 map units from B (Punnett, 1940). Telloni et al. (1976) studied the t(OH 10), a translocation between the Z and a microchromosome, and found that the K and dw loci were located on the arm that contained the break point. The B locus showed no linkage with the break point. Blazak and Fechheimer (1980) conducted a C-band analysis of this same translocation and reported that the terminal C-band (G-band light staining region) on the Z chromosome was located on the intact arm. Kaelbling and Fechheimer (1983b) obtained measurements of the synaptonemal complex
n
Chi-square 1.01
recovered, showing that extremely tight genetic linkage exists between these markers. Data for the NM 7659 X id+ (Z-linked dermal melanin) mating are presented in Table 4. There was no significant linkage relationship shown between these markers.
TABLE 4. Test for linkage of the NhI 7659 with Z-linked dermal melanin
t(Z;l)
Gamete type
n
Chi-square
T-IdO
58
XT=1.67
U-id+0
54
4 r -60 xL-l-07
1.66
N-Id
56
217.0***
T-id+
72 240
Parental gametic type; T = rearrangement carrier, N = normal karyotype.
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linkage relationships found between y and either B or S. The data for the MN t(Z;l) X y backcross mating are shown in Table 2. A significant linkage of 18.2 ± 3.2 map units was calculated between these markers. The data for the MN t(Z;3) X B-S backcross mating are presented in Table 3. As described in the Materials and Methods section, the numbers differ between the B and the S analyses, because black chicks from the first two hatches were not reared to determine segregation at the S locus. Independent assortment was noted between the break point and B. No recombinants between the break point and S were
Z CHROMOSOME LINKAGES
arm opposite the arm containing the terminal G-band light staining region, again showing that the S-K-dw region is on the p arm of the Z chromosome. This MN (Z;3)-S intimate relationship is the third interchange/morphological trait relationship of this type that has been found in the chicken. The first reported was the NM 7092 translocation (Zartman, 1973), which showed complete linkage to a protoporphyrin mutant (pr) (Shoffner et al, 1982). The second was complete linkage of a pericentric inversion within chromosome 2 with a limb development mutation that was "locked into" the inverted segment (Langhorst and Fechheimer, 1983). Specific regions of three chromosomes have now been defined by both cytological and morphological markers. The definition of the By region of the q arm of the Z chromosome will be useful for further linkage studies. These two easily identifiable markers are now mapped in relation to a common, almost central marker (the MN t(Z;l)), and have extended the sweep distance for genetic testing of this region of the q arm to greater than 50 map units. The id+ locus, known to be linked to B by 10 map units, will be tested against the MN t(Z;l) to attempt to determine its location within this chromosome region. Also, F t males have been obtained that are genotypically id+-B-y/Id-b+-Y+ in order to attempt to further refine linkage relationships between Id, B, and Y+ in this region. Results from this mating should also disclose whether or not y will interact with id+ in a manner similar to the interaction of id+ and autosomal dominant white skin (W+), which results in blue or slaty blue/black shanks (Hutt, 1949). While independence was noted between B and y, use of the MN t(Z;l) interchange showed that these loci are in fact located on the same arm of the Z chromosome, another indication of the apparently extremely long genetic distances in the larger chicken chromosomes. Punnett (1940) also discussed this in a review of data gathered by himself and several other investigators. At that time, the possibility that the largest chromosome was the sex chromosome was being discussed. It is now known that the Z chromosome is the fifth largest in the chicken, and its genetic length is apparently longer than the average length of any chromosome of the laboratory mouse (Womack, 1979). This reinforces the importance of utilizing
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formed by this t(OH 10) rearrangement and determined that the arm that was involved in the rearrangement was the short arm (Fechheimer, personal communication). The results of these four studies indicate that the G-band light staining region of the Z chromosome marks the long (q) arm of this chromosome. The break point on the Z chromosome in the MN t(Z;l) is in the G-band light staining region (Bitgood, 1980). This break point, therefore, is on the q arm of the Z chromosome. Previous mapping of this same rearrangement against B showed 22 map units between these markers (Bitgood etal, 1980). The linkage of 18 map units shown between this break point and y (Table 2) and the independence found between B and y (Table 1), suggest the linear order of £(22)T(18)y (T representing the translocation break point) on the long arm of the Z chromosome. Females hemizygous for B-y have been mated to homozygous MN t(Z;l) males to obtain F! males that will be used in a three point test cross to verify these findings. It is not yet possible to determine which direction the centromere or the telomere lie from this chromosome segment. Significant deviations from expected numbers of Y+ and y individuals are noted in Table 2. Sixty-two individuals were scored as y and 97 were scored as Y+. In the same table, there is also a deviation from expected in the segregation of the translocation, 50 normals and 84 translocation carriers. These are possibly chance deviations, as other matings using either the translocation or y have segregated normally (unpublished observations). The results from the future test mating involving the MN t(Z;l), B, and y (described above) will be monitored to determine if any unusual segregations might be occurring in this chromosomal region. A contingency chi-square for linkage was calculated using the procedure presented by Green (1963). From the results of the MN t(Z;3) X B, S backcross mating, it is evident that this break point is genetically very closely linked to S. Due to the reduction in crossing over expected in the vicinity of an interchange, it is not known what the physical distance between these markers might be. Further tests of this rearrangement against other loci on the short (p) arm of the chromosome will clarify this relationship. Wang and Shoffner (1974) conducted G-band analysis of this interchange and demonstrated that the break point was on the
2237
2238
BITGOOD
multiple point test crosses in chicken gene linkage studies in order to define regions for expansion of sweep distances for additional linkage tests. ACKNOWLEDGMENTS
REFERENCES Bitgood, J. J., 1980. Gametogenesis and zygote formation in domestic fowl (Gallus domesticus) with chromosome rearrangements. Ph.D. Thesis, University of Minnesota, St. Paul, MN. Bitgood, J. J., R. N. Shoffner, J. S. Otis, and W. E. Briles, 1980. Mapping of the genes for pea comb, blue egg, barring, silver, and blood groups A, E, H, and P in the domestic fowl. Poultry Sci. 59:1686- 1693. Blazak, W. F., and N. S. Fechheimer, 1980. Gonosome-autosome translocations in fowl: meiotic configurations and chiasma counts from singly and doubly heterozygous cockerals. Can. J. Genet. Cytol. 22:343-351.. Campo, J. L., and F. Orozco, 1980. The action of the sex linked barring gene on Spanish chickens with gold plumage. Ann. Genet. Sel. Anim. 12: 233-239. Green, M. C , 1963. Methods for testing linkage. Pages 56—82 in Methodology in Mammalian Genetics. W. J. Burdette, ed. Holden-Day, Inc., San Francisco, CA. Hutt, F. B., 1949. Genetics of the Fowl. McGraw-Hill Book Company, Inc., New York, NY. Kaelbling, M., and N. S. Fechheimer, 1983a. Synaptonemal complexes and the chromosome complement of domestic fowl, Gallus domesticus. Cytogenet. Cell Genet. 35:87-92.
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Appreciation is expressed to R. N. Shoffner, University of Minnesota and N. S. Fechheimer, The Ohio State University for furnishing the chromosome rearrangement lines used in this study. Appreciation is also expressed to L. Neumann, W. Schumann, E. Eutsler, and B. Rhiner for technical assistance.
Kaelbling, M., and N. S. Fechheimer, 1983b. Synaptonemal complex analysis of chromosome rearrangements in domestic fowl, Gallus domesticus. Cytogenet. Cell Genet. 36:567— 572. Langhorst, L. J., and N. S. Fechheimer, 1983. Correlation between presence of a pericentric inversion of the second chromosome and limb phenotype in the domestic chicken. J. Dairy Sci. 66 (Suppl. 1):246. (Abstr.) McGibbon, W. H., 1981. White skin: a Z-linked recessive mutation in the fowl. J. Hered. 72: 139-140. Punnett, R. C , 1940. Genetic studies in poultry. X. Linkage data for the sex chromosome. J. Genet. 39:335-342. Shoffner, R. N., A. Krishan, G. J. Haiden, R. K. Bammi, and J. S. Otis, 1967. Avian chromosome methodology. Poultry Sci. 56:334—344. Shoffner, R. N., R. Shuman, J. S. Otis, J . J . Bitgood, V. Garwood, and P. Lowe, 1982. The effect of a protoporphyrin mutant on some economic traits of the chicken. Poultry Sci. 61:817-820. Somes, R. G., Jr., 1984. Linked loci of the chicken — Gallus gallus (G. domesticus). Genet. Maps 3:465-473. Telloni, R. V., R. G. Jaap, and N. S. Fechheimer, 1976. Cytogenetic and phenotypic effects of a chromosomal rearrangement involving the Zchromosome and a micro-chromosome in the chicken. Poultry Sci. 55:1886-1896. Wang, N., and R. N. Shoffner, 1974. Trypsin G- and C-banding for interchange analysis and sex identification in the chicken. Chromosoma (Berl.) 47:61-69. Wang, N., R. N. Shoffner, J. S. Otis, and K. M. Cheng, 1982. The induction of chromosomal structural changes in male chickens by the alkylating agents: triethylene melamine and ethyl methanesulfonate. Mutat. Res. 96:53—66. Womack, J. E., 1979. Linkage map: mouse. Pages 38—39 in Inbred and Genetically Defined Strains of Laboratory Animals. Part 1. Mouse and Rat. P. L. Altman and D. D. Katz, ed. Fed. Am. Soc. Exper. Biol., Bethesda, MD. Zartman, D. L., 1973. Location of the pea comb gene. Poultry Sci. 52:1455-1462.