Showing papers on "Ribosomal DNA published in 1983"
TL;DR: The Xenopus 28S rDNA has nine major areas of sequence inserted when compared to E. coli 23S r DNA, and in one insert there are 25 bases completely conserved between the various eukaryotes, suggesting that this area is important for eUKaryotic ribosomes.
Abstract: We have determined the complete nucleotide sequence of Xenopus laevis 28S rDNA (4110 bp). In order to locate evolutionarily conserved regions within rDNA, we compared the Xenopus 28S sequence to homologous rDNA sequences from yeast, Physarum, and E. coli. Numerous regions of sequence homology are dispersed throughout the entire length of rDNA from all four organisms. These conserved regions have a higher A + T base composition than the remainder of the rDNA. The Xenopus 28S rDNA has nine major areas of sequence inserted when compared to E. coli 23S rDNA. The total base composition of these inserts in Xenopus is 83% G + C, and is generally responsible for the high (66%) G + C content of Xenopus 28S rDNA as a whole. Although the length of the inserted sequences varies, the inserts are found in the same relative positions in yeast 26S, Physarum 26S, and Xenopus 28S rDNAs. In one insert there are 25 bases completely conserved between the various eukaryotes, suggesting that this area is important for eukaryotic ribosomes. The other inserts differ in sequence between species and may or may not play a functional role.
165 citations
TL;DR: It is shown here that the level of transcription of the injected ribosomal genes is strongly affected by spacer sequences far upstream of the promoter, and it is proposed that the upstreamSpacer sequences act to influence the frequency of promoter activation.
130 citations
TL;DR: Results suggest that protein C23 has a preference for binding DNA sequences in the nontranscribed spacer of rDNA.
Abstract: Protein C23, a predominant nucleolar phosphoprotein and a putative nucleolus organizer protein, was analyzed for its general DNA binding characteristics and for its selectivity in binding plasmid DNAs containing cloned fragments of the genes that code for ribosomal RNA (rDNA). By use of nitrocellulose filter disk assays, the protein bound saturably to nuclear DNA with a relatively high affinity. Binding was maximal at low ionic strength (0-0.1 M KCl) with progressively decreasing binding at or above 0.2 M. In competition assays protein C23 showed a marked preference for linear single-stranded vs. double-stranded DNA and little or no affinity for ribosomal RNA. The relative affinities of rDNA sequences for protein C23 were determined with cloned fragments spanning 15.8 kilobases (kb) of DNA starting approximately 3.7 kb upstream from the initiation site for 45S preribosomal RNA to near the 3' end of the sequence coding for 28S RNA. Of the five linearized plasmids tested, only one (pKW1) was an effective competitor for 32P-labeled nuclear DNA. As measured by the concentration of competing DNA required to achieve 50% competition, pKW1 was approximately 20-fold more effective than the second best competitor. The DNA insert in pKW1 is a 3.5-kb sequence which is located in the nontranscribed spacer region less than 0.5 kb upstream from the initiation site for 45S preribosomal RNA. These results suggest that protein C23 has a preference for binding DNA sequences in the nontranscribed spacer of rDNA.
103 citations
TL;DR: The four ribosomal RNA genes of the rodent malaria parasite were identified and mapped by restriction enzyme analysis and Southern blot hybridization of genomic DNA and restriction analysis confirms both the low copy number and that these genes are not in an easily recognizable tandem array.
84 citations
TL;DR: Ribosomal DNA fragments from the human malaria parasite Plasmodium falciparum have been cloned and analysed in detail and copy number analysis reveals that each fragment is represented approximately 4 times in the genome, and implies that there are a total of 8 rRNA genes organised into at least two classes of transcription unit.
Abstract: Ribosomal DNA fragments from the human malaria parasite Plasmodium falciparum have been cloned and analysed in detail. Restriction mapping shows that the cloned fragments are different. However, they do have some similarities, in particular a small stretch of A+T-rich DNA located between the small and large subunit rRNA genes. A small rRNA gene has been mapped to this A+T-rich region. Copy number analysis reveals that each fragment is represented approximately 4 times in the genome, and implies that there are a total of 8 rRNA genes organised into at least two classes of transcription unit. Analysis of a third overlapping rDNA fragment indicates that the large subunit rRNA gene of at least one transcription unit contains an intervening sequence.
62 citations
TL;DR: XC cell line was established from a rat fibrosarcoma induced by avian sarcoma virus and analysis of ribosomal RNA genes in the DNA of this cell line indicated that there was a 3- to 4-fold increase in the number of the rDNA repeating units.
62 citations
TL;DR: The region of chromosome XII containing theRDN1 gene which encodes the cytosolic ribosomal RNA molecules and the region of chromosomes III containing theHIS4 (histidine 4) gene were analysed in 30 lager yeast strains, 11 ale strains and 20 strains from a number of different species in the genus Saccharomyces.
Abstract: The region of chromosome XII containing theRDN1 gene which encodes the cytosolic ribosomal RNA molecules and the region of chromosome III containing theHIS4 (histidine 4) gene were analysed in 30 lager yeast strains, 11 ale strains and 20 strains from a number of different species in the genus Saccharomyces.
61 citations
TL;DR: The nucleotide sequences of the two internal transcribed spacers, the adjacent ribosomal coding sequences and the boundary between the external transcribed spacer and the 18S coding sequence in a cloned ribosome transcription unit from Xenopus borealis are determined.
Abstract: We have determined the nucleotide sequences of the two internal transcribed spacers, the adjacent ribosomal coding sequences and the boundary between the external transcribed spacer and the 18S coding sequence in a cloned ribosomal transcription unit from Xenopus borealis. The transcribed spacers differ very extensively from those of X. laevis. Nevertheless, embedded in the internal transcribed spacers are several short sequence elements which are identical between the two species. These conserved elements are laterally displaced by substantial distances in the X. borealis sequence with respect to that of X. laevis. These relative displacements imply that insertions and deletions have played a major role in transcribed spacer divergence in Xenopus. This in turn implies that large regions of the transcribed spacers do not play a sequence-specific role in ribosome maturation. In contrast, the sequenced parts of the ribosomal coding regions, which encompass 670 nucleotides, differ at only three points from the corresponding sequences in X. laevis, each by a single substitution. These substitutions are readily accommodated by current models for rRNA higher order structure.
59 citations
TL;DR: The complete nucleotide sequence of a 23S rRNA gene from a blue-green alga, Anacystis nidulans, has been determined and has strong homology with the chloroplast 4.5S r RNA.
55 citations
TL;DR: The polarity of transcription of the rDNA was determined by hybridizing 3'-OH end labeled rRNA to Southern blots of genomic and cloned rDNA and the results indicate that a transcription unit of P. berghei rDNA is arranged as in other organisms with the small rRNA at the 5' end and the large r RNA at the 3' end.
55 citations
TL;DR: In this paper, the structure of ribosomal chromatin in differentiating cells was examined using essentially the same methods, and the combined results suggest that whereas regions that are not transcribed are packaged with proteins that protect them from nuclease digestion, actively transcribing riboomal genes are associated with few macromolecular constituents apart from those required for transcription and its regulation.
TL;DR: In situ hybridization of this probe to metaphase chromosomes demonstrates that rDNA and CAD genes do coamplify in two independently isolated PALA-resistant mutants, consistent with the model that unequal exchanges between rDNA genes mediate the amplification of CAD genes in the Syrian hamster mutants that were analyzed.
Abstract: The amplified CAD genes in N-(phosphonacetyl)-L-aspartate (PALA)-resistant Syrian hamster cells are located in an expanded chromosomal region emanating from the site of the wild-type gene at the tip of the short arm of chromosome B-9. The terminus of B-9 in PALA-sensitive cells contains a cluster of rRNA genes (i.e., a nucleolus organizer, rDNA). We have used a molecular clone containing sequences complementary to Syrian hamster 28S rRNA to investigate whether rDNA is coamplified with CAD genes in the PALA-resistant mutants. In situ hybridization of this probe to metaphase chromosomes demonstrates that rDNA and CAD genes do coamplify in two independently isolated PALA-resistant mutants. The tight linkage of CAD and rDNA genes was demonstrated by their coordinate translocation from B-9 to the end of the long arm of chromosome C-11 in one mutant. Blot hybridization studies substantiate the in situ hybridization results. Both types of analysis indicate that only one or two rDNA genes, on the average, are coamplified per CAD gene. The data are consistent with the model that unequal exchanges between rDNA genes mediate the amplification of CAD genes in the Syrian hamster mutants that were analyzed.
TL;DR: Although no stems analogous to those bracketing 16S and 23S rRNA in Escherichia coli pre-rRNA are evident, certain features of the spacer regions flanking eucaryotic mature rRNAs are conserved and could be involved in rRNA processing or ribosome formation.
Abstract: Two regions of mouse rDNA were sequenced. One contained the last 323 nucleotides of the external transcribed spacer and the first 595 nucleotides of 18S rRNA; the other spanned the entire internal transcribed spacer and included the 3' end of 18S rRNA, 5.8S rRNA, and the 5' end of 28S rRNA. The mature rRNA sequences are very highly conserved from yeast to mouse (unit evolutionary period, the time required for a 1% divergence of sequence, was 30 X 10(6) to 100 X 10(6) years). In 18S rRNA, at least some of the evolutionary expansion and increase in G + C content is due to a progressive accretion of discrete G + C-rich insertions. Spacer sequence comparisons between mouse and rat rRNA reveal much more extensive and frequent insertions and substitutions of G + C-rich segments. As a result, spacers conserve overall G + C richness but not sequence (UEP, 0.3 X 10(6) years) or specific base-paired stems. Although no stems analogous to those bracketing 16S and 23S rRNA in Escherichia coli pre-rRNA are evident, certain features of the spacer regions flanking eucaryotic mature rRNAs are conserved and could be involved in rRNA processing or ribosome formation. These conserved regions include some short homologous sequence patterns and closely spaced direct repeats.
TL;DR: The ribosomal DNA of Cucurbita pepo L. has been found to consist of tandemly arrayed repeat units, most of which are 10 kilobases in length, which differ from each other in length and/or nucleotide sequence.
Abstract: The ribosomal DNA (rDNA) of Cucurbita pepo L. has been found to consist of tandemly arrayed repeat units, most of which are 10 kilobases in length. Thirty-six repeat units, cloned into the HindIII site of pACYC 177, fall into seven classes which differ from each other in length and/or nucleotide sequence. Most of the heterogeneity occurs in noncoding portions of the repeat unit although there is some nucleotide sequence variation in the coding portion as well. Heterogeneity of base modification was observed in genomic rDNA of which two examples are: (a) all of the repeat units have three BamHI sites, one of which is unavailable for restriction in about half of the units and (b) all of the CCGG sites except one are methylated at the internal cytidine in many of the units; a second site is unmethylated in some of the units and in a very few units a third site remains unmethylated.
TL;DR: The transcription of flax rDNA and the processing of the rRNA precursors is described and one of the least methylated regions maps close to the 5' end of the largest identified rRNA precursor.
Abstract: The transcription of flax rDNA and the processing of the rRNA precursors is described. The location of methylated regions of the rDNA has been assayed, one of the least methylated regions maps close to the 5' end of the largest identified rRNA precursor.
TL;DR: Selective transcription of hybrid plasmids containing yeast rDNA was achieved with a template-dependent S100 fraction from whole cell extracts of Saccharomyces cerevisiae and RNA polymerase I traverses the interspersed 5 S rRNA gene.
TL;DR: The structure and number of the ribosomal RNA (rRNA) genes of the avian malaria parasite Plasmodium lophurae has been examined using Southern blot analysis and recombinant DNA techniques and it is revealed that there are approximately six rRNA genes per haploid genome.
Abstract: The structure and number of the ribosomal RNA (rRNA) genes of the avian malaria parasite Plasmodium lophurae has been examined using Southern blot analysis and recombinant DNA techniques. The ribosomal DNA (rDNA) of P. lophurae has been cloned into the plasmid pBR322, beginning with size-selected populations of Cla I- and Hind III-restricted parasite DNA. The structure of two clones (CL-1 and HA-2) is presented in detail. These two clones together probably represent the entire 17s and 25s coding regions of P. lophurae. Analysis of quantitative genomic Southern blots reveals that there are approximately six rRNA genes per haploid genome, and that the rRNA genes may be divided into four distinct classes by restriction analysis. Examination of the flanking regions of these genes indicates that they are not organized into easily recognizable tandem repeats.
TL;DR: The X.borealis external transcribed spacer is some 612 nucleotides long, about 100 nucleotide shorter than in X.laevis.
Abstract: We report sequence data from a cloned rDNA unit from Xenopus borealis, extending leftwards from the 18S gene to overlap a region previously sequenced by R. Bach, B. Allet and M. Crippa (Nucleic Acids Research 9, 5311-5330). Comparison with data from other species of Xenopus leads to the inference that the transcription initiation site in X.borealis is in the newly sequenced region and not, as was previously thought, in the region sequenced earlier. The X.borealis external transcribed spacer thus defined is some 612 nucleotides long, about 100 nucleotides shorter than in X.laevis. The X.borealis and X.laevis external transcribed spacers show a pattern of extensive but interrupted sequence divergence, with a large conserved tract starting about 100 nucleotides downstream from the transcription initiation site and shorter conserved tracts elsewhere. The regions in between the conserved tracts differ in length between the respective external transcribed spacers indicating that insertions and deletions have contributed to their divergence, as previously inferred for the internal transcribed spacers. Much of the overall length difference is in the region flanking the 18S gene, where there are also length microheterogeneities in X.laevis rDNA. As in X.laevis, the transcribed spacer sequences flanking the 18S gene in X.borealis contain no major tracts of mutual complementarity. The accumulated data on transcribed spacers in Xenopus render it unlikely that processing of ribosomal precursor RNA involves interaction between the regions flanking 18S RNA.
TL;DR: The restriction endonuclease cleavage pattern of the rDNA from an X chromosome undergoing magnification is examined, and there is no evidence for the selective amplification of either uninterrupted rDNA units or those containing insertion sequences.
TL;DR: The large ribosomal repeat of the Lupinus luteus genome is 8.8 kb in length as revealed by Southern blotting and hybridization to 5.8 S rRNA and cloned rDNA fragments of L. luteUS and Schizosaccharomyces pombe.
TL;DR: The recombination and repair defective mutant, mei-41, exhibits three rather striking effects on the genetic properties and chromosomal stability of rDNA in Drosophila, and these effects are explained in a simple model that has general implications for chromosome structure.
Abstract: The recombination and repair defective mutant, mei-41 , exhibits three rather striking effects on the genetic properties and chromosomal stability of rDNA in Drosophila. First, mei-41 inhibits rDNA magnification. However, mei-9 , another recombination and repair defective mutation has no similar effect. This indicates that magnification requires some, but not all, of the gene products necessary for meiotic exchange. Second, under magnifying conditions, mei-41 induces interchanges between the X rDNA and either arm of the Ybb - chromosome. These interchanges occur at high frequency and are independent of rDNA orientation. Third, in mei-41 bb +/ Ybb + males, bobbed mutants in the X , but not the Y , also arise at high frequency. Evidence suggests that these events involve the rDNA type I insertion. The recombination and repair defective properties of mei-41 together with our results regarding its unusual and specific effects involving rDNA are explained in a simple model that has general implications for chromosome structure.
TL;DR: Ribosomal DNA from Xenopus laevis contains several heterogeneities in all three transcribed spacers, as revealed by analysis of cloned and uncloned amplified rDNA from oocytes and cloned chromosomal r DNA from erythrocytes.
Abstract: Ribosomal DNA (rDNA) from Xenopus laevis contains several heterogeneities in all three transcribed spacers, as revealed by analysis of cloned and uncloned amplified rDNA from oocytes and cloned chromosomal rDNA from erythrocytes. Heterogeneities include single base changes and length variants of one to several nucleotides. Sites of variation are widely but non-uniformly distributed, some occurring only a short distance outside the boundaries of the rRNA coding regions. No two transcription units that we have yet examined are identical throughout their transcribed spacer regions.
TL;DR: Comparison of the mouse and human ribosomal DNA transcription initiation sites with the rat sequence reported in this paper demonstrates a conserved sequence at +2 to +16, CTGACACGCTGTCCT, which suggests that this region may be important for the initiation of transcription on mammalian Ribosomal DNAs.
Abstract: The transcription initiation site for rat 45S precursor ribosomal RNA synthesis was determined by nuclease protection mapping with two single-strand endonucleases. S1 and mung bean, and one single-strand exonuclease, ExoVII. These experiments were performed with end-labeled ribosomal DNA from double-stranded pBR322 recombinants and from single-stranded M13 recombinants. Results from experiments using both kinds of DNA and all three enzymes showed that the 5' end of 45S RNA mapped to a unique site 125 bases upstream from the Hind III site in the ribosomal DNA gene. The DNA surrounding this site (designated +1) was sequenced from -281 to +641. The entire sequence of this region shows extensive homology to the comparable region of mouse. This includes three stretches of T residues in the non-coding strand between +300 and +630. Two sets of direct repeats adjacent to these T-rich regions are observed. Comparison of the mouse and human ribosomal DNA transcription initiation sites with the rat sequence reported in this paper demonstrates a conserved sequence at +2 to +16, CTGACACGCTGTCCT. This suggests that this region may be important for the initiation of transcription on mammalian ribosomal DNAs.
TL;DR: A DNAseI hypersensitive site in the ribosomal DNA spacer of Xenopus laevis and Xenopus borealis is detected, and it is concluded that this domain plays an important role in rDNA function.
Abstract: We have detected a DNAseI hypersensitive site in the ribosomal DNA spacer of Xenopus laevis and Xenopus borealis. The site is present in blood and embryonic nuclei of each species. In interspecies hybrids, however, the site is absent in unexpressed borealis rDNA, but is present normally in expressed laevis rDNA. Hypersensitive sites are located well upstream (over lkb) of the pre-ribosomal RNA promoter. Sequencing of the hypersensitive region in borealis rDNA, however, shows extensive homology with the promoter sequence, and with the hypersensitive region in X. laevis. Of two promoter-like duplications in each spacer, only the most upstream copy is associated with hypersensitivity to DNAaseI. Unlike DNAaseI, Endo R. MspI digests the rDNA of laevis blood nuclei at a domain extending downstream from the hypersensitive site to near the 40S promoter. Since the organisation of conserved sequence elements within this "proximal domain" is similar in three Xenopus species whose spacers have otherwise evolved rapidly, we conclude that this domain plays an important role in rDNA function.
TL;DR: The reason for the interest in this system is that rDNA exemplifies both the intriguing and the perplexing aspects of DNA methylat ion.
Abstract: Like many cellular macromolecules, DNA is subject tc'postsynthetic modification. In animals, the most common modif icat ion involves methylat ion of cytosine in the dinucleotide sequence CpG (Wyatt 1951; Doscocil and Sorm 1962; Grippo et al. 1968; Vanyushin et al. 1970). Current interest in CpG methylation arises from three lcinds of observations. First, the presence or absence of cytosine methylation determines the interaction of DNA with restriction endonucleases in bacteria (Meselson et al. 1972; Mann and Smith 1978), and this has encouraged speculation that protein-DNA interact ions are similarly control led in eukaryotes (Holl iday and Pugh 1975, Riggs 1975). Second, there is evidence that the pattern of DNA methylation in a cell can be inherited by i ts daughter cel ls (for review, see Wigler l98l). The idea of a heri table, but reversible, switch on the DNA has obvious attract ions. Final ly, t issuespecific alterations in the methylation pattern of several genes have been observed. [n general, the level of methylation at a gene is lower in cells transcribing the sequence than in nontranscribing cells (see, e.g., Kuo et al. 1979; Mandel and Chambon 1979; van der Ploeg and Flavel l 1980; Bird et a l . l98lb; Weintraub et a l . l98l ; for review, see Doerf ler 198 1 ; Ehrl ich and Wang 198 1). In some cases i t has been demonstrated direct ly that transcription is inhibited when the DNA to be tested is first methylated in vitro (Vardimon et al. 1982). Our recent research on the function of DNA methylat ion has concentrated on the genes for 28S and l8S ribosomal RNAs (rRNAs). In animals, the r ibosomal DNA (rDNA) is made up of uni ts compris ing a nontranscribed spacer, plus a transcribed region that codes for the rRNA precursor. Units are tandemly repeated at one or more sites in the genome (for review, see Lewin 1980). The reason for our interest in this system is that rDNA exemplifies both the intriguing and the perplexing aspects of DNA methylat ion. Thus, in dif ferent organisms, rDNA can be found unmethylated (e.g., invertebrates), heavily methylated (e.9., amphibia), or a combination of both (e.g., mouse)(Rae and Steele l9l9; Bird and Taggart 1980; Bird et al. 1981a). Any model for the function of DNA methylation must explain this variability. In spite of the range of methylation patterns, however, there is good correspondence between expression of the r ibosomal genes and general or local ized absence of methylation. For example, in BALB/c mice the unmethylated fraction of rDNA is preferentially sensit ive to DNase I in isolated l iver nuclei (Bird et al. l98la). This impl ies that the methylated fract ion of
TL;DR: Analysis of Physarum amoebal strains showed that rDNA was inherited in an all-or-nothing fashion; that is, progeny clones contained one or the other parental rDNA type, but not both; one r DNA type was inherited more frequently than the other.
Abstract: In the acellular slime mold Physarum polycephalum, the several hundred genes coding for rRNA are located on linear extrachromosomal DNA molecules of a discrete size, 60 kilobases. Each molecule contains two genes that are arranged in a palindromic fashion and separated by a central spacer region. We investigated how rDNA is inherited after meiosis. Two Physarum amoebal strains, each with an rDNA recognizable by its restriction endonuclease cleavage pattern, were mated, the resulting diploid plasmodium was induced to sporulate, and haploid progeny clones were isolated from the germinated spores. The type of rDNA in each was analyzed by blotting hybridization, with cloned rDNA sequences used as probes. This analysis showed that rDNA was inherited in an all-or-nothing fashion; that is, progeny clones contained one or the other parental rDNA type, but not both. However, the rDNA did not segregate in a simple Mendelian way; one rDNA type was inherited more frequently than the other. The same rDNA type was also in excess in the diploid plasmodium before meiosis, and the relative proportions of the two rDNAs changed after continued plasmodial growth. The proportion of the two rDNA types in the population of progeny clones reflected the proportion in the parent plasmodium before meoisis. The rDNAs in many of the progeny clones contained specific deletions of some of the inverted repeat sequences at the central palindromic symmetry axis. To explain the pattern of inheritance of Physarum rDNA, we postulate that a single copy of rDNA is inserted into each spore or is selectively replicated after meiosis.
TL;DR: The principal rDNA repeating unit in the MitchellPetunia genome has a length of 8.5 kb; however, additional sequence changes have occurred since the ‘short’ repeat is characterized by restriction endonuclease cleavage sites which are absent in the longer variant units.
Abstract: The principal rDNA repeating unit in the MitchellPetunia genome has a length of 8.5 kb. In addition there is a major variant of length 9.7 kb, and two minor variants of 9.3 kb and 10.4 kb. The size heterogeneity of the rDNA repeating units results from length differences in the non-transcribed spacer regions. These differences may reflect simple insertions into the non-transcribed spacer region of the major 'short' repeat; however, additional sequence changes have occurred since the 'short' repeat is characterized by restriction endonuclease cleavage sites which are absent in the longer variant units.
TL;DR: Results from S-1 endonuclease analysis suggest that the s-16S rDNA region is probably not transcribed into stable s- 16S rRNA, and a structural model of the "truncated rRNA operon" is presented.
Abstract: An extra 16S rRNA gene (s-16S rDNA) from the Euglena gracilis chloroplast genome and several hundred positions of its flanking regions have been sequenced. The structural part has 1486 positions and is to 98% homologous in its sequence with the 16S rRNA gene in functional chloroplast rRNA operons. Sequences of about 200 positions upstream and 15 positions downstream of the structural part of the s-16S rRNA gene region are highly homologous with corresponding parts in the functional operon. Neither tRNA genes (A1a, I1e) nor parts of the 23S and 5S rRNA genes are found within 557 positions after the 3' end of the s-16S rRNA gene, i.e., the 330 bp homology, observed in electron microscopic studies of heteroduplexes (4), between the s-16S rDNA downstream region and the 6.2 kb repeated segment containing the functional rRNA operon, must be due to a DNA stretch in the interoperon spacer. A structural model of the "truncated rRNA operon" is presented. Results from S-1 endonuclease analysis suggest that the s-16S rDNA region is probably not transcribed into stable s-16S rRNA.
TL;DR: This observation suggests that gross deletions of rDNA can result in nucleolus organizer regions with predominantly one type of repeat unit, in agreement with the observation that In+ repeats do not significantly contribute to functional rRNA.
Abstract: The nucleolus organizer (NO) of the D. melanogaster X chromosome is composed of ribosomal repeat units which contain two types (I and II) of non-rDNA insertions (In+) and repeats with no insertions (In-). Evidence from other laboratories indicate random interspersion of all types of repeat units within the X NO. An EcoRI and BamHI examination of rDNA from two bobbed mutants, bb2rl and mal12 demonstrates segregation of the major type I repeat units. The 46 rDNA repeats of the bb2rl NO contain no detectable major type I repeats whereas the majority of the 68 rDNA mal12 repeats are major type I and tandemly linked. This observation suggests that gross deletions of rDNA can result in nucleolus organizer regions with predominantly one type of repeat unit. Additivity tests demonstrate that the 46 ribosomal repeats of the bb2rl chromosome revert the phenotype of other bobbed NOs, but the 68 mal12 ribosomal repeats show no or slight additivity. This is in agreement with the observation that In+ repeats do not significantly contribute to functional rRNA. A Southern blot analysis using BamHI which cuts only in type I insertions demonstrates that the majority of major type I In+ repeating units exist in tandem linkage group(s) within the X NO.