Abstract
Interband DNA of Drosophila melanogaster polytene chromosomes was studied using a novel approach based on the electron microscopic (EM) analysis of chromosome regions carrying DNA fragements of known molecular genetic composition, inserted by P element-mediated transformation. Insertion of such fragments predominantly into interbands makes it possible to clone interband DNA by constructing genomic libraries from transformed strains and probing them with the insert DNA. The transformed strain P[H-sp70:Adh](61C) has insertion in the 61 C7-8 interband on the left arm of chromosome 3. This DNA consists of part of the hsp70 gene promoter fused to the coding region of the Adh gene, and is flanked on either side by P element sequences. We constructed a genomic library from DNA of this strain and isolated a clone containing the insert and the interband DNA. Subsequently the genomic library of wild-type strain was probed with a subclone composed of interband DNA only. We have thus isolated a clone containing the entire native interband. 1289 by of interband DNA was sequenced and found to be AT-rich (53.4%) with numerous regions of overlapping direct and inverted repeats, regulatory sites, and two overlapping open reading frames (ORFs).
Similar content being viewed by others
References
Ashburner M (1989) Drosophila. A laboratory handbook. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York
Beermann W (1972) Chromosomes and genes. In: Beermann W (ed) Results and problems in cell differentiation. Vol 4, Springer-Verlag, Berlin-Heidelberg-New York, pp 1–13
Bonner JJ, Parks C, Parker-Thornberg J, Mortin MA, Pelham HRB (1984) The use of promoter fusions in Drosophila genetics: isolation of mutations affecting the heat shock response. Cell 37:979–991
Crick F (1971) General model for the chromosomes of higher organisms. Nature 234:25–27
Gall JG, Pardue ML (1971) Nucleic acid hybridization in cytological preparations. Methods Enzymol 21:470–480
Goldberg DA (1980) Isolation and partial characterization of the Drosophila alcohol dehydrogenase gene. Proc Natl Acad Sci USA 77:5794–5798
Jamrich M, Greenleaf AL, Bautz EK (1977) Localization of RNA polymerase in polytene chromosomes of Drosophila melanogaster. Proc Natl Acad Sci USA 74:2079–2083
Keppy DO, Welshons WJ (1977) The cytogenetics of a recessive visible mutant associated with a deficiency adjacent to the Notch locus in Drosophila melanogaster. Genetics 85:497–506
Kerkis AY, Zhimulev IF, Belyaeva ES (1977) EM autoradiographic study of 3H-uridine incorporation into Drosophila melanogaster salivary gland chromosomes. Drosophila Inf Serv 52:14–17
Kidd S, Hockett TS, Young MW (1983) The Notch locus of Drosophila melanogaster. Cell 34:421–433
Laski FA, Rio DC, Rubin GM (1986) Tissue specificity of Drosophila P element transposition is regulated at the level of mRNA splicing. Cell 44:7–19
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning. A laboratory manual Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York
Maxam AM, Gilbert W (1980) Sequences end-labeled DNA with base-specific chemical cleavages. Methods Enzymol 65:499–560
Mott MR, Burnett EJ, Hill RJ (1980) Ultrastructure of polytene chromosomes of Drosophila isolated by microdissection. J Cell Sci 45:15–30
Mott MR, Hill RJ (1986) The ultrastructural morphology of native salivary gland chromosomes of Drosophila melanogaster: The band — interband question. Chromosoma 94:403–411
O'Hare K, Rubin GM (1983) Structures of P transposable elements and their sites of insertion and excision in the Drosophila melanogaster genome. Cell 34:25–35
O'Kane CJ, Gehring WS (1987) Detection in situ of genomic regulatory elements in Drosophila. Proc Natl Acad Sci USA 84:9123–9127
Paul J (1972) General theory of chromosome structure and gene activation in eukaryotes. Nature 238:444–446
Pirrotta V (1986) Cloning Drosophila genes. In: Roberts DB (ed) Drosophila: A practical approach. IRL Press, Oxford-Washington DC, pp 83–110
Ramos RGP, Grimwade BG, Wharton KA, Scottgale TN, Artavanis-Tsakonas S (1989) Physical and functional definition of the Drosophila Notch locus by P element transformation. Genetics 123:337–348
Rubin GM, Spradling AC (1983) Vector P element-mediated gene transfer in Drosophila. Nucleic Acids Res 11:6341–6351
Rykowski MC, Parmelee SJ, Agard DA, Sedat JW (1988) Precise determination of the molecular limits of a polytene chromosome band: regulatory sequences for the Notch gene are in the interband. Cell 54:461–472
Sass H (1982) RNA polymerase B in polytene chromosomes: immunofluorescent and autoradiographic analysis during stimulated and repressed RNA synthesis. Cell 28:269–278
Sass H, Bautz EK (1982a) Immunoelectron microscopic localization of RNA polymerase B on isolated polytene chromosomes of Chironomus tentans. Chromosoma 85:633–642
Sass H, Bautz EK (1982b) Interbands of polytene chromosomes: binding sites and start points for RNA polymerase B (II). Chromosoma 86:77–93
Semeshin VF, Zhimulev IF, Belyaeva ES (1979) Electron microscope autoradiographic study on transcriptional activity of Drosophila melanogaster polytene chromosomes. Chromosoma 73:163–177
Semeshin VF, Baricheva EM, Belyaeva ES, Zhimulev IF (1982) Electron microscopical analysis of Drosophila polytene chromosomes. I: Mapping of the 87A and 87C heat shock puffs in development. Chromosoma 87:229–237
Semeshin VF, Belyaeva ES, Zhimulev IF, Lis JT, Richards G, Bourouis M (1986) Electron microscopical analysis of Drosophila polytene chromosomes. IV. Mapping of morphological structures appearing as a rasult of transformation of DNA sequences into chromosomes. Chromosoma 93:461–468
Semeshin VF, Demakov SA, Perez Alonso M, Belyaeva ES, Bonner JJ, Zhimulev IF (1989) Electron microscopical analysis of Drosophila polytene chromosomes. V. Characteristics of structures formed by transposed DNA segments of mobile elements. Chromosoma 97:396–412
Skaer RJ (1977) Interband transcription in Drosophila. J Cell Sci 26:251–266
Solovyev VV, Rogozin IB (1986) The “CONTEXT” package of computer programs for contextual analysis of DNA, RNA and protein sequences. I. Searching for sequence similarities and functional sites. Institute of Cytology and Genetics Publication Novosibirsk
Sorsa V (1975) A hypothesis for the origin and evolution of chromomere DNA. Hereditas 81:77–84
Spradling AC, Rubin GM (1982) Transposition of cloned P elements into Drosophila germ line chromosomes. Science 218:341–347
Vlassova IE, Umbetova GH, Zimmermann VH, Alonso JA, Belyaeva ES, Zhimulev IF (1985) Immunofluorescence localization of DNA:RNA hybrids in Drosophila melanogaster polytene chromosomes. Chromosoma 91:251–258
Whitmore T (1986) Localization of a “housekeeping” gene in Chironomus thummi thummi. Cytobios 46:193–200
Zhimulev IF, Belyaeva ES, (1975) 3H-uridine labeling patterns in the Drosophila melanogaster salivary gland chromosomes X, 2R, 3L. Chromosoma 49:219–231
Zhimulev IF, Belyaeva ES, Semeshin VF (1981) Informational content of polytene chromosome bands and puffs. Crit Rev Biochem 11:303–340
Author information
Authors and Affiliations
Additional information
Communicated by A. Ashburner
Rights and permissions
About this article
Cite this article
Demakov, S.A., Semeshin, V.F. & Zhimulev, I.F. Cloning and molecular genetic analysis of Drosopbila melanogaster interband DNA. Molec. Gen. Genet. 238, 437–443 (1993). https://doi.org/10.1007/BF00292003
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00292003