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. Author manuscript; available in PMC: 2009 Sep 28.
Published in final edited form as: Cell. 1989 Mar 24;56(6):957–968. doi: 10.1016/0092-8674(89)90629-6

Figure 3. Identification of Mutations in the Cyclin A Gene.

Figure 3

(A) Cytogenetic map of the region on chromosome arm 3L containing the cyclin A gene. The regions deleted in the chromosomal deficiencies vin2 and vin3 (Akam et al., 1978) are indicated by black bars. In situ hybridization localized the cyclin A sequence within the two proximal breakpoints of vin2 and vin3 (arrow). Five lethal complementation groups (rsg11–rsg15) have been localized between these breakpoints (Hoogwerf et al., 1988). The EMS-induced allele I(3)183 of the complementation group rsg11 complements neither the EMS-induced allele I(3)v4-4 nor the allele neo114 isolated after P element insertion mutagenesis (Cooley et al., 1988)

(B) Mapping the P element insertion neo114. Southern analysis with a cyclin A cDNA probe was used to compare genomic restriction fragments from an rsg11 allele (neo114), a deletion of the region (vin3), and a control chromosome. The chromosome carrying the allele neo116 was chosen as control because it is derived from the same parental chromosome as neo114 and it also has a P element insert but in a different location. Since the chromosomes being compared carry recessive lethals, DNA was prepared from heterozygous flies. Novel fragments absent from the control DNA (neo116/TM3, lanes 1, 3, and 7) but apparent in neo114/TM3 DNA are marked with arrowheads. Lanes 1 and 2: Sall; lanes 3 and 4: EcoRI; lanes 5–8: HindIII. Despite the appearance of novel fragments, no loss of bands is observed in Sall and EcoRI digests of neo114/TM3 DNA because of the contribution of the balancer chromosome. However, one band present in the control (lane 7) cannot be detected in HindIII digests of neo114/TM3 DNA (lane 8, arrow). This disappearance indicates the presence of a restriction site polymorphism. In digests of DNA from flies carrying a deficiency deleting the cyclin A gene (vin3), the hybridizing fragments are uniquely derived from the balancer chromosome, and a comparison of the banding pattern derived from two different balancer chromosomes, In(3L)P (lane 5) and TM3 (lane 6), reveals a HindIII restriction site polymorphism. This polymorphism allows the unambiguos assignment of the hybridizing fragments to the different chromosomes present in neo116/TM3 flies or neo114/TM3 flies, respectively. Position and size (kbp) of molecular weight markers are indicated on the left side.