Fig. 2

Molecular-genetic mapping of the unplugged locus to LG 10. (A) Schematic drawing showing the three-generation mapping cross. In the G₀ generation, a heterozygous unplugged fish in the Tü background was crossed to a polymorphic WIK fish. F₁ heterozygous fish were crossed to each other, F₂ homozygous mutants and their siblings were collected, and their DNA was extracted for chromosomal mapping. (B) PCR amplification using marker Z3835 and DNA isolated from the G₀ Tü fish, the G₀ WIK fish, pooled F₂ mutants, and pooled F₂ siblings. The Tü -specific PCR fragment is shorter than the WIK-specific fragment. Amplification with Z3835 in F₂ sibling pool DNA results in a Tü -specific and a WIK-specific fragment, while in DNA from pooled mutant only the Tü -specific fragment is present (higher mobility), suggesting linkage between Z3835 and the unplugged mutation. (C) PCR amplification using marker Z3835 and DNA from individual F₂ sibling embryos. Embryos in which only a WIK-specific fragment is present (lower mobility) are of +/+ genotype, while embryos with a Tü -specific and a WIK-specific PCR fragment are heterozygotes (+/- genotype). (D) PCR amplification using marker Z3835 and DNA from individual F₂ unplugged mutant embryos. In most mutant embryos, only the Tü -specific fragment is present. In recombinant embryos, a Tü -specific and a WIK-specific fragment are present (*). In these embryos, the unplugged Tü chromosome recombined with the WIK chromosome between Z3835 and the unplugged locus. (E) Schematic representation of LG 10 around the unplugged locus. The centromere is indicated by a black circle.