Linkage Analysis and Mapping of Three Sex-Linked Color Pattern Genes in the Guppy, Poecilia reticulata

Gideon Khoo, Tit Meng Lim, Woon-Khiong Chan and Violet P. E. Phang*

As pointed out by Khoo et. al. (1999) many exotic strains have been developed in the last few decades by “intensive selection of spontaneous mutant genes that affect the coloration as well as the shape and size of the body and fins (Dzwillo, 1959; Kirpichnikov, 1981; Fernando and Phang, 1985).” This intensive selection, which continues at present days, is responsible for all past, present and future strains.

In this article we summarize a scientific publication about the genetic linkage of three sex-linked color pattern genes: black caudal-peduncle ( Bcp), red tail ( Rdt) and variegated tail ( Var) (Khoo et al., 1999a, b), their interactions with each other, and their relationships to the blue tail ( Blt), green tail ( Grt) and snakeskin body-snakeskin tail ( Ssb-Sst) traits that were investigated in our earlier studies (See genetic map below). Understanding how these are linked is important to the breeder working with these traits.

Remember that:

• Almost all the genes involved in color pigmentation and patterning are sex-linked.
• It has 23pairs of chromosomes, 22 of which are autosomal and one the sex chromosomes.
• Male guppies are heterogametic (XY) while the females are homogametic (XX).
• Genes for background body coloration, e.g., blond ( b), gold ( g), albino ( a) and blue ( bl) are, however, autosomally inherited and recessive to their wild-type alleles.
• Color patterns on the body and fins of domesticated guppy strains take the form of single colors, snakeskin-like reticulations and variegated mosaic patterns of two or more colors.
• Expression of phenotypic color patterns in cultured guppies has been found to be determined by dominant sex-linked and sex-limited genes.
• Consequently, these genes may be used as genetic (phenotypic) markers to map the X- and Y chromosomes of the guppy.

ORIGINAL BROODSTOCK FOR TEST CROSSES IN THIS STUDY

Three- to four-week old fry of the Tuxedo (TUX) and Green Variegated (GV) guppy strains were obtained from highly inbred and well established TUX and GV stocks of the Chin Lam Brothers Tropical Fish Farm and Swee Hing & Brothers Aquarium Co., respectively, in Singapore. Tuxedo and Green Variegated are the commercial names given to these strains by guppy breeders. Male and female juveniles, distinguishable by the expression of their color patterns due to sexual dimorphism, were cultured separately according to Khoo et al. (1999a, b) for another three to four weeks before being used for reciprocal crosses between the TUX and GV strains. This was to ensure that juvenile males were fully mature (as indicated by a well developed gonopodium) and females had not been previously inseminated. Under laboratory conditions, domesticated guppies reach sexual maturation at six to eight weeks of age.

Fig. 1. (A) Adult male guppy of the Tuxedo (TUX) strain showing a black caudal-peduncle and red tail. (B) Adult female guppy of the TUX strain with grey caudal-peduncle and faint red tinges on an opaque greyish-white tail. (C) Adult male guppy of the Green Variegated (GV) strain displaying an orange tail with yellow streaks, and numerous black spots and patterns of different shapes and sizes. (D) Adult female guppy of the GV strain with wild-type female olive-brown background body coloration and faint greyish-brown variegated patterns on a yellowish tail.

RECIPROCAL CROSSES

To establish the mode of inheritance and linkage of the black caudal-peduncle, red tail and variegated tail color patterns, single-pair reciprocal crosses were made between six-week old mature virgin fish of the TUX and GV strains. Singlepair full-sib F1 males and F1 females were mated to produce the F2 generation. Newly born fry were separated and raised to maturity in 3.5-liter clear plastic tanks (five fish/tank). F1 and F2 offspring were segregated according to color phenotype and sex.

RESULTS

Segregation and recombination in F1 and F2 offspring of TUX x GV

Nine mating pairs of TUX x GV produced a total of 132 male and 131 female F1 offspring (Table 1A). F1 males had the black caudal-peduncle and red caudal fin of their TUX male parents but also displayed black spots and patches on the tail fin (Fig. 2A). F1 females had a grey caudal-peduncle and an opaque greyish-white tail with red tinges and black spots (Fig. 2B).

Fig. 2. Progenies (F1, F2 and recombinants) from Tuxedo x Green Variegated and Green Variegated x Tuxedo crosses had the following phenotypic color patterns for males and females…

Fig. 2(A). males: (a) Tuxedo (TUX), (b) Tuxedo with variegated tail patterning (TUXVAR), (c) red tail with variegated patterns (RTVAR), (d) black caudal-peduncle with variegated tail patterns (BCPVAR), (e) red tail (RT) and (f) variegated tail with a mosaic pattern of large black spots and patches (VAR).

Fig. 2(B). Females: (a) TUXVAR, (b) RTVAR, (c) VAR and (d) TUX.

DISCUSION

From these crosses, Khoo et al, (1999) demonstrated that “these color patterns are simple sex-linked traits controlled by single genes”. In addition, Khoo et al, (1999) show that “Bcp, Rdt and Var are dominantly expressed in both males and females, albeit the colors are more distinct and definitive in the males due to the presence of androgens (Figs. 1, 2)”

Each of the three color pattern genes has two alleles: Bcp which is dominant over Bcp+, Rdt dominant over Rdt+ and Var over Var+. Recessive alleles of these loci do not give rise to any color patterns.

This study proves that the Bcp, Rdt and Var genes are able to cross over from the X-chromosome to the Y and vice versa since male and female recombinants of the TUX, VAR, RT, TUXVAR, RTVAR and BCPVAR phenotypes were observed at the F2

Scientists have showed that the “X- and Y-chromosomes of the guppy are equal in size and indistinguishable by ordinary cytological methods. As a result of this homology between the sex chromosomes, genes are able to crossover along almost the whole length of their chromatids. Only a small segment on the Y-chromosome, the sex-determining region (SdR) which is presumed to contain male-determining genes, is known to be non-homologous and different from the X.”

CONCLUSION

The black caudal-peduncle ( Bcp), red tail ( Rdt) and variegated tail ( Var) color pattern genes of the domesticated guppy are

1. single genes located at three different different loci,
2. dominantly expressed,
3. X- and Y-linked, and
4. fully capable of crossing-over from the Y- to the X-chromosome and vice versa.

Map distances for sex-determining region (SdR)– Rdt, Rdt–Bcp, SdR– Bcp and SdR– Var are approximately 3.1, 2.3, 5.1 and 2.2 map units, respectively, with a gene order of Var–SdR– Rdt–Bcp.