A Genetic Analysis of the rose - gespbten Region ( 6 SCS - 69 BS ) of Drosophila melanogaster

We describe a genetic analysis of the region 68C8-6YB5 defined by D[(3L)uin-7. We have induced 35 new lethal mutations in this region, which together with 20 existing lethal mutations, visible mutations, genes identified by protein products and one gene deduced from complementation data fall into 37 complementation groups in this %-band interval. Using existing and newly induced deficiencies we have assigned these to 11 intervals defined by deficiency breakpoints. Those mutations which fell in the same breakpoint interval as the Lsp-2 gene, which codes for the abundant larval serum protein 2, were the subject of detailed study. None was rescued by the active h p 2 gene transformed on to chromosome I1 and we conclude that, as yet, we have n o lethal mutations Of Lsp-2. W E report here the isolation and complementation mapping of lethal mutations in the region 68C8-69B5, on the left arm of chromosome ZZZ of Drosophila melanogmter, as defined by Df(3L)vin-7. This project was initiated to provide a better map of the region of the Lsp-2 gene, which encodes the second major protein component of the serum of late third instar larvae (ROBERTS, WOLFE and AKAM 1977), and in an attempt to isolate recessive lethal mutations of that gene. Our hope was to learn something of the function of this abundant protein from the phenotype of Lsp-2 mutations. In this we have failed. The results define five lethal complementation groups lying between the deficiency breakpoints that flank the Lsp-2 locus, but we reject all of these as candidates for Lsp-2 mutations. The mutations recovered do however define 32 lethal complementation groups, in addition to the 3 visible and 2 protein coding loci located within this 35-band region. MATERIALS AND METHODS Culture conditions: Stocks were maintained on standard yeast-cornmeal-agar medium in half-pint milk bottles at 25". Nipagin was added as an inhibitor of moulds. For cytological studies and for the analysis of hemolymph proteins flies were reared on yeast-glucose-agar medium (10% yeast, 10% glucose and 3% agar autoclaved after mixing). The temperature-sensitive stocks were kept at 18" and transferred to 29" as necessary. Stocks: The Oregon-R strain maintained in this laboratory for over 15 yr was used as the reference wild-type ' Present address: Department of Medical Genetics, Churchill Hospital, Present address: Genetics Department, University ofCambridge, DownHeadington, Oxford, England. ing Street, Cambridge CB2 3EH, England. ' To whom reprint requests should be sent. Genetics 118 665-670 (April, 1988). strain. Unless indicated otherwise the LSP-2 of all stocks was indistinguishable from that of the Oregon-R stock. Many of the stocks used in this work have been described by us previously AKAM (1977), AKAM, ROBERTS and W01.w (1978) and AKAM et al. (1978). Additional stocks were obtained as follows: Tp(3 :3)LIbx"'"/TM 1 Me (Tp(3: 3)68A68E;89E), E. B. LEWIS, California Institute of Technology; 1(3)V11-5 p ' eITMJ(ln(3LR) TM3 Sb Sere) A. St IEAKN, Johns Hopkins University, Baltimore; and lethal mutants in the Lsp-2 region (Table 1) from E. MEYEKOWI.I.Z, California Institute of Technology. The Ightham stocks (pronounced "item") were established from a gravid female caught in the village of Ightham in Kent (AKAM el al. 1978). The remaining stocks have been in this laboratory for many years and the phenotypes of all these mutants are described in LINUSLEY and GRELL (3968) and LINDSLEY and ZIMM (1985, 1986, 1987) with the exception of uin (ANXOIAHEI~EKE and PERIQUEL. 1973). One allele of every complementation group described here will be maintained by us for reference purposes for at least '12 months and will be deposited at the European Stock Centre in Ume%. EMS mutagenesis: Males homozygous for the markers uin and apP, which flank the Lsp-2 locus and marked with u (which in the presence of uin gives a bright orange eye) were mutagenized by the method of LEWIS and BACHER (1968). Lethal mutations were recovered as shown in Figure 1. Complementation: All lethal mutations were balanced over TM3 and crossed to deficiencies of the Lsp-2 region (Table 2), which were also balanced over TM3. The absence of Sb' flies in more than 100 progeny was taken as the criterion for the failure to complement. By this technique mutations were placed between deficiency breakpoints. Where two lethals occurred between the same two deficiency breakpoints they were crossed inter se. The absence of Sb+ flies in more than 100 progeny was again taken as the criterion for the failure to complement. Nomenclature of mutations: The region of 3L which we study here is flanked distally by rs (rose) and proximally by gs (gespleten). Following the precedent of JUDD, SHEN 666 A. M. Hoogwerf, M. Akam and D. Roberts