Four core genotypes mouse model: localization of the Sry transgene and bioassay for testicular hormone levels - PubMed (original) (raw)

Four core genotypes mouse model: localization of the Sry transgene and bioassay for testicular hormone levels

Yuichiro Itoh et al. BMC Res Notes. 2015.

Abstract

Background: The "four core genotypes" (FCG) mouse model has emerged as a major model testing if sex differences in phenotypes are caused by sex chromosome complement (XX vs. XY) or gonadal hormones or both. The model involves deletion of the testis-determining gene Sry from the Y chromosome and insertion of an Sry transgene onto an autosome. It produces XX and XY mice with testes, and XX and XY mice with ovaries, so that XX and XY mice with the same type of gonad can be compared to assess phenotypic effects of sex chromosome complement in cells and tissues.

Findings: We used PCR to amplify the Sry transgene and adjacent genomic sequences, to resolve the location of the Sry transgene to chromosome 3 and confirmed this location by fluorescence in situ hybridization (FISH) of the Sry construct to metaphase chromosomes. Using quantitative PCR, we estimate that 12-14 copies of the transgene were inserted. The anogenital distance (AGD) of FCG pups at 27-29 days after birth was not different in XX vs. XY males, or XX vs. XY females, suggesting that differences between XX and XY mice with the same type of gonad are not caused by difference in prenatal androgen levels.

Conclusion: The Sry transgene in FCG mice is present in multiple copies at one locus on chromosome 3, which does not interrupt known genes. XX and XY mice with the same type of gonad do not show evidence of different androgen levels prenatally.

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Figures

Figure 1

Figure 1

Location of the Sry transgene in four core genotypes mouse model. Samples were from XX and XY mice with testes (XX_Sry_, XY− Sry) and XX and XY mice with ovaries (XX, XY−). Confirmation of transgene-genome boundary by PCR using transgene-specific primer d and Chr3 primer e (A) , with transgene-specific primer c and Chr3 primer b (B), and with transgene-specific primer c and Chr3 primer a (C). (D) The concatemer of Sry transgene is inserted into a repetitive sequence present genome-wide. (E) Sry transgene location on chromosome 3. (F) A visual estimate of the difference in copy number of Sry between wildtype and XY− Sry genomic DNA in agarose gel.

Figure 2

Figure 2

FISH mapping of the Sry transgene to Chromosome 3. The Sry transgene signal (A, red) and chromosome Chromosome 3 paint (B, green) hybridize to the same metaphase chromosome. (C) Sry transgene hybridization in three additional metaphase cells demonstrating its location with respect to the p- and q-arm ends.

Figure 3

Figure 3

Anogenital distance measurement of FCG mice. Asterisk shows the significant effect of sex (two way ANOVA, p < 0.00001).

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