Summary
Restriction and hybridization analysis of cucumber native ribosomal (r) DNA purified from actinomycin-D/CsCl gradients suggested that the repeat units were heterogeneous in both length and sequence. Several full length rDNA repeat units were cloned and five are described which account for all the EcoR I and Xba I fragments present in native DNA. One of a number of BamH I sites found in the clones is not found in a proportion of native rDNA because of base modification. Restriction maps are described for the representative clones and aligned with R-loop maps obtained from electron microscope analysis of each type of repeat unit hybridized under R-loop conditions to pure 18S and 25S rRNAs. The major heterogeneity is explained by differences in length of the external spacer region and by a proportion of the repeat units showing a restriction fragment length polymorphism on EcoR I digestion. The regions coding for 18S and 25S rRNA are uninterrupted and highly conserved.
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References
Brown DD, Gurdon JB (1964) Absence of ribosomal RNA synthesis in the anucleolate mutant of X. laevis. Proc Natl Acad Sci USA 51:139–146
Chaconas G, Sande JH van de (1980) 5′-32P labelling of RNA and DNA restriction fragments. Methods Enzymol 65:75–85
Chang ACY, Cohen SN (1978) Construction and characterization of amplifiable multicopy cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol 134:1141–1156
Davis RW, Simon M, Davidson N (1971) Electron microscope heteroduplex methods for mapping regions of base sequence homology in nucleic acids. Methods Enzymol 21D:413–428
Domoney C, Timmis JN (1980) Ribosomal RNA gene redundancy in juvenile and mature ivy (Hedera helix). J Exp Bot 31:1093–1110
Dover GA, Flavell RB (1984) Molecular coevolution: DNA divergence and the maintenance of function. Cell 38:622–623
Duggleby RG, Kimms H, Rood JI (1981) A computer programme for determining the size of DNA restriction fragments. Anal Biochem 110:49–55
Ellis THN, Davies DR, Castleton JA, Bedford ID (1984) The organization and genetics of rDNA length variants in peas. Chromosoma 91:74–81
Gerlach WL, Bedbrook JR (1979) Cloning and characterization of ribosomal RNA genes from wheat and barley. Nucleic Acids Res 7:1869–1885
Goldsbrough PB, Cullis CA (1981) Characterization of the genes for ribosomal RNA in flax. Nucleic Acids Res 9:1301–1309
Gross-Bellard M, Oudet P, Chambon P (1973) Isolation of high molecular weight DNA from mammalian cells. Eur J Biochem 36:32–38
Grunstein M, Hogness D (1975) Colony hybridization. A method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci USA 72:3961–3965
Hemleben V, Grierson D, Dertman H (1977) The use of equilibrium centrifugation in actinomycin-caesium chloride for the purification of ribosomal DNA. Plant Sci Lett 9:129–135
Ingle J, Timmis JN, Sinclair J (1975) The relationship between satellite DNA, ribosomal RNA gene redundancy and genome size in plants. Plant Physiol 55:496–501
Locker J (1979) Analytical and preparative electrophoresis of RNA in agarose-urea. Anal Biochem 98:358–367
Maggini F, Carmona MJ (1981) Sequence heterogeneity of the ribosomal DNA in Allium cepa (Liliaceae). Protoplasma 108:163–171
Maizels N (1976) Dictyostelium 17S, 25S and 5S rDNAs lie within a 38,000 base pair repeated unit. Cell 9:431–438
Maniatis T, Hardison RC, Lacy E, Laner J, O'Connell C, Quon D, Skin GK, Efstratiadis A (1978) The isolation of structural genes from libraries of eukaryotic DNA. Cell 15:687–701
Matsuda K, Siegel A (1967) Hybridization of plant ribosomal RNA to DNA — the isolation of a DNA component rich in ribosomal RNA cistrons. Proc Natl Acad Sci USA 58:673–680
McClelland M (1983) The effect of site specific methylation on restriction endonuclease cleavage (update). Nucleic Acids Res 11:169–173
Murray NE, Brammer WJ, Murray K (1977) Lamboid phages that simplify the recovery of in vitro recombinants. Mol Gen Genet 150:53–61
Peacock AC, Dingham CW (1967) Resolution of multiple ribonucleic acid species by polyacrylamide gel electrophoresis. Biochemistry 6:1818–1827
Rafalski JA, Wiewiorowski M, Söll D (1983) Organization of ribosomal DNA in yellow lupine (Lupinus luteus) and sequence of the 5.8S gene. FEBS Lett 152:241–245
Reeder RH (1984) Enhancers and ribosomal gene spacers. Cell 38:349–351
Rozek CE, Timberlake WE (1979) Restriction endonuclease mapping by crossed-contact hybridization: the ribosomal RNA genes of Achlya ambisexualis. Nucleic Acids Res 7:1657–1678
Scott NS, Ingle J (1973) The genes for cytoplasmic RNA in higher plants. Plant Physiol 51:677–684
Siegel A, Kolacz K (1983) Heterogeneity of pumpkin ribosomal DNA. Plant Physiol 72:166–171
Southern E (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517
Thomas M, White RL, Davis RW (1976) Hybridization of RNA to double stranded DNA: formation of R-loops. Proc Natl Acad Sci USA 73:2294–2298
Timmis JN, Sinclair J, Ingle J (1972) Ribosomal RNA genes in euploids of hyacinth. Cell Differ 1:335–339
Tullis RH, Rubin H (1980) Calcium protects DNase I from proteinase K: a new method for the removal of contaminating RNase I from DNase. Anal Biochem 107:260–264
Varsanyi-Breiner A, Gusella JF, Keys C, Housman DE, Sullivan D, Brisson N, Verma DPS (1979) The organization of a nuclear DNA sequence from a higher plant: molecular cloning and characterization of soybean ribosomal DNA. Gene 7:317–334
Waldron J, Dunsmuir P, Bedbrook J (1983) Characterization of the rDNA repeat units in the Mitchell petunia genome. Plant Mol Biol 2:57–65
Wellauer PK, Dawid IB (1979) Isolation and sequence organization of human ribosomal DNA. J Mol Biol 128:289–303
Yakura K, Kato A, Tunfuji S (1983) Structural organization of ribosomal DNA in four Trillium species and Paris verticillata. Plant Cell Physiol 24:1231–1240
Yakura K, Kato A, Tanifuji S (1984) Length heterogeneity of the larger spacer of Vicia faba is due to the differing number of a 325 bp repetitive sequence. Mol Gen Genet 193:400–405
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Communicated by D. von Wettstein
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Kavanagh, T.A., Timmis, J.N. Heterogeneity in cucumber ribosomal DNA. Theoret. Appl. Genetics 72, 337–345 (1986). https://doi.org/10.1007/BF00288570
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DOI: https://doi.org/10.1007/BF00288570