Simple Sequence Motifs Research Articles

The unusually long small-subunit ribosomal RNA of the crustacean, Daphnia pulex: sequence and predicted secondary structure.

The small-subunit (SSU) ribosomal RNA (rRNA) gene of the Branchiopod crustacean, Daphnia pulex, was isolated from a phage library and subcloned into the plasmid pBluescript. Determination of the complete nucleotide sequence of this gene revealed it to be 2293 nt in length with a G + C content of 53.7%. This is the longest SSU rRNA gene yet reported from a crustacean. The predicted secondary structure of the rRNA is quite typical for eukaryotes except for length expansion in four regions that are known to be highly variable: V2, V4, V7, and V9. Increases in V4 and V7 were most notable. RT-PCR analysis of these two variable regions showed that they are present in the mature rRNA molecule. Potential secondary structures for these regions are proposed based on energetic criteria. Sequence simplicity analysis of V4 and V7 did not reveal the occurrence of clustered simple sequence motifs. This suggests that replication slippage may not be an important mechanism generating the length expansion in these regions as has been proposed for unusually long regions in insect SSU rRNA genes. On the other hand, there was a pronounced bias in nucleotide composition in the variable regions (G + C = 56.6% in V4 and 60% in V7) which is typical of long SSU rRNA genes in insects. Comparison of the lengths of regions V2, V4, V7, and V9 among diverse arthropods revealed that substantial increases in V4 and V7 tend to co-occur. This suggests the possibility of a functional relationship between these variable regions.

Molecular structure of the rDNA intergenic spacer (IGS) in Triturus: implications for the hypervariability of rDNA loci.

Ribosomal DNA (rDNA) variation in the species Triturus vulgaris meridionalis (Amphibia, Urodela) is remarkable because of unusually high intraspecific variability in the number and distribution of ribosomal loci in the karyotype; in addition, portions of the intergenic spacer (IGS) are clustered at chromosomal loci where they are not associated with ribosomal 18S and 28S RNA genes. These clusters are referred to as extraribosomal, and they appear to consist mostly of repetitive BamHI elements. In this paper, we report the complete nucleotide sequence of an IGS of T. v. meridionalis; this structural analysis is aimed to get insight into the molecular mechanism(s) of spreading of the ribosomal cistrons as well as its possible functional significance. We found that the IGS of T. vulgaris has a modular structure: modular repetitive elements contain sequences possibly related to the regulation of transcription of the ribosomal units. In particular, both ribosomal and extraribosomal IGS elements contain presumptive enhancers. Interestingly, the enhancer-containing region is mostly conserved between ribosomal and extraribosomal elements, while mutations accumulate in a region characterized by repetitions of a simple sequence motif, that we consider as a possible recombination hotspot. Our data suggest that extraribosomal elements most probably originated from ribosomal enhancer-containing elements able to move independently from the ribosomal unit at novel chromosomal positions, perhaps with the aid of the simple repetitive motif. We argue that a similar mechanism may lead to the spreading of complete repetition units as well, giving rise to multiple, and variable, ribosomal sites. We propose that hypervariability in the number and distribution of the rDNA loci, as seen in T. vulgaris, is a further mechanism to ensure redundancy, which seems to be an intrinsic property of rDNA biology, the occurrence of IGS elements independently clustered at separate chromosomal loci being a by-product of this mechanism.

Isolation and characterization of microsatellite loci from the planarian Dugesia polychroa (Schmidt) (Platyhelminthes:Tricladida).

Molecular EcologyVolume 6, Issue 4 p. 389-391 Isolation and characterization of microsatellite loci from the planarian Dugesia polychroa (Schmidt) (Platyhelminthes: Tricladida) S. Ramachandran, S. Ramachandran Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorL. W. Beukeboom, L. W. Beukeboom Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorL. Gerace, L. Gerace Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorN. Pavlovic, N. Pavlovic Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorS. Carranza, S. Carranza Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorN. K. Michiels, N. K. Michiels Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this author S. Ramachandran, S. Ramachandran Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorL. W. Beukeboom, L. W. Beukeboom Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorL. Gerace, L. Gerace Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorN. Pavlovic, N. Pavlovic Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorS. Carranza, S. Carranza Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this authorN. K. Michiels, N. K. Michiels Arbeitsgruppe Michiels, Max-Planck-Institut für Verhaltensphysiologie, D-82319 Seewiesen, GermanySearch for more papers by this author First published: 31 October 2003 https://doi.org/10.1046/j.1365-294X.1997.00190.xCitations: 4 Dr L. Beukeboom Tel.: +49-8157-932254 or 232; Fax: +49-8157 932400 or -209. E-mail: [email protected] AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL No abstract is available for this article.Citing Literature Volume6, Issue4April 1997Pages 389-391 RelatedInformation

Abundance and length polymorphism of microsatellite repeats in Beta vulgaris L.

Simple sequence repeats (SSRs) are known to exhibit high degrees of variability even among closely related individuals. Their usage as nuclear genetic markers requires their conversion into sequence-tagged sites (STSs). In this paper we present the development of simple sequences as STSs for Beta vulgaris. This species comprises wild, cultivated, and weedy forms; the latter are thought to originate from accidental hybridisation between the other two. Two partial genomic libraries were screened with simple sequence motifs (AT, CA, CT, ATT, GTG, and CA, CT, respectively). Clones of 22 CA, nine CT, eight ATT, and one GTG sequence were obtained. AT micro satellites were present in compound motifs, not recognised by the probe. Sequence comparisons revealed that 20 CA clones containing short motifs (<16 bp) were variants of a previously described approximately 320-bp satellite DNA (Schmidt et al. 1991), and hence did not correspond to unique loci. Polymorphism of one (ATT)15 and three (CT)n, with n=15, 17 and 26, was detected by PCR on a sample of 64 plants from the different forms of B. vulgaris. 13 (ATT), 13 (CT), nine (CT) alleles and one (CT) allele were detected. One of the ATT alleles was much larger than the others (>800 bp). Genetic variability was high among wild beets, lower among cultivated beets, and intermediate among weed beets. One allele of each locus was found at high frequencies in cultivated beets and, to a lower extent, in weed beets. The combination of three polymorphic loci allowed the individual identification of 17/17 wild and 15/15 weed beets, and 21/32, mostly homozygous, cultivated beets.

Northern blot analysis of simple repetitive sequence transcription in plants

The presence of simple repetitive sequence motifs in RNA from various plant species was probed by Northern blot analysis. Hybridization of total, poly(A)(+)- and poly(A)(-)-RNA with microsatellite-complementary oligonucleotide probes revealed distinct bands with most but not all probe/species combinations, demonstrating the presence of di-, tri- and tetranucleotide repeat motifs in plant transcripts. Only trinucleotide repeat-derived hybridization signals were found to be enriched in the poly(A)(+)-fraction. The quality of Northern blot signals proved to be highly dependent on hybridization stringency. Thus, under the stringency conditions usually applied for oligonucleotide hybridization, some probes [(GT)8, (CAC)5, (TCC)5, and (CCTA)4] cross-hybridized to bands corresponding in size to 18S and/or 26S rRNA. Cross-hybridization to rRNA was significantly reduced at higher stringencies. These results stress the importance of carefully adjusting the hybridization conditions in Northern blot analysis of simple sequence transcripts.

The contribution of slippage-like processes to genome evolution

Simple sequences present in long (> 30 kb) sequences representative of the single-copy genome of five species (Homo sapiens, Caenorhabditis elegans, Saccharomyces cerevisiae, E. coli, and Mycobacterium leprae) have been analyzed. A close relationship was observed between genome size and the overall level of sequence repetition. This suggested that the incorporation of simple sequences had accompanied increases of genome size during evolution. Densities of simple sequence motifs were higher in noncoding regions than in coding regions in eukaryotes but not in eubacteria. All five genomes showed very biased frequency distributions of simple sequence motifs in all species, particularly in eukaryotes where AAA and TTT predominated. Interspecific comparisons showed that noncoding sequences in eukaryotes showed highly significantly similar frequency distributions of simple sequence motifs but this was not true of coding sequences. ANOVA of the frequency distributions of simple sequence motifs indicated strong contributions from motif base composition and repeat unit length, but much of the variation remained unexplained by these parameters. The sequence composition of simple sequences therefore appears to reflect both underlying sequence biases in slippage-like processes and the action of selection. Frequency distributions of simple sequence motifs in coding sequences correlated weakly or not at all with those in noncoding sequences. Selection on coding sequences to eliminate undesirable sequences may therefore have been strong, particularly in the human lineage.

Abundance and polymorphism of di-, tri-and tetra-nucleotide tandem repeats in chickpea (Cicer arietinum L.)

The abundance and polymorphism of 38 different simple-sequence repeat motifs was studied in four accessions of cultivated chickpea (Cicer arietinum L.) by in-gel hybridization of synthetic oligonucleotides to genomic DNA digested with 14 different restriction enzymes. Among 38 probes tested, 35 yielded detectable hybridization signals. The abundance and level of polymorphism of the target sequences varied considerably. The probes fell into three broad categories: (1) probes yielding distinct, polymorphic banding patterns; (2) probes yielding distinct, monomorphic banding patterns, and (3) probes yielding blurred patterns, or diffused bands superimposed on a high in lane background. No obvious correlation existed between abundance, fingerprint quality, and the sequence characteristics of a particular motif. Digestion with methyl-sensitive enzymes revealed that simple-sequence motifs are enriched in highly methylated genomic regions. The high level of intraspecific polymorphism detected by oligonucleotide fingerprinting suggests the suitability of simple-sequence repeat probes as molecular markers for genome mapping.

How is it that microsatellites and random oligonucleotides uncover DNA fingerprint patterns?

Minisatellites, microsatellites, and short random oligonucleotides all uncover highly polymorphic DNA fingerprint patterns in Southern analysis of genomic DNA that has been digested with a restriction enzyme having a 4-bp specificity. The polymorphic nature of the fragments is attributed to tandem repeat number variation of embedded minisatellite sequences. This explains why DNA fingerprint fragments are uncovered by minisatellite probes, but does not explain how it is that they are also uncovered by microsatellite and random oligonucleotide probes. To clarify this phenomenon, we sequenced a large bovine genomic BamHI restriction fragment hybridizing to the Jeffreys 33.6 minisatellite probe and consisting of small and large Sau3A-resistant subfragments. The large Sau3A subfragment was found to have a complex architecture, consisting of two different minisatellites, flanked and separated by stretches of unique DNA. The three unique sequences were characterized by sequence simplicity, that is, a higher than chance occurrence of tandem or dispersed repetition of simple sequence motifs. This complex repetitive structure explains the absence of Sau3A restriction sites in the large Sau3A subfragment, yet provides this subfragment with the ability to hybridize to a variety of probe sequences. It is proposed that a large class of interspersed tracts sharing this complex yet simplified sequence structure is found in the genome. Each such tract would have a broad ability to hybridize to a variety of probes, yet would exhibit a dearth of restriction sites. For each restriction enzyme having 4-bp specificity, a subclass of such tracts, completely lacking the corresponding restriction sites, will be present. On digestion with the given restriction enzyme, each such tract would form a large fragment.(ABSTRACT TRUNCATED AT 250 WORDS)

Mammalian telomere dynamics: healing, fragmentation shortening and stabilization

Telomeres are essential for stable chromosome maintenance. The simple G-rich sequence motif d(TTAGGG)n is all that is required in cis for telomere function in mammalian cells, as in other eukaryotes. Using this fact, telomeres have been used to specifically fragment mammalian chromosomes to dissect their structure and function. Telomere length maintenance is altered in cancer cells. Trans-acting factors, such as telomerase and telomere-binding proteins, may determine telomere function in both normal and cancer cells. Current experiments are aimed at understanding the role of telomerase and telomere-binding proteins in cellular senescence and immortalization.

Oligonucleotide fingerprinting of isolates of Candida species other than C. albicans and of atypical Candida species from human immunodeficiency virus-positive and AIDS patients

Oligonucleotide fingerprinting of genomic DNA from oral isolates of four different Candida species other than C. albicans and atypical chlamydospore-positive isolates from human immunodeficiency virus (HIV)-positive individuals and AIDS patients was investigated as a means for differentiating between isolates within individual species. Oligonucleotides composed of simple repetitive sequence motifs, including (GACA)4, (GATA)4, (GGAT)4, (GTG)5, and (GT)8, all yielded fingerprints suitable for strain segregation of 8 C. tropicalis isolates, 12 Torulopsis (Candida) glabrata isolates, 8 atypical Candida isolates, and, except for (GATA)4, 2 C. krusei probe in turn and so generate several distinct DNA fingerprints of the same DNA sample. However, none of the probes yielded fingerprints suitable for strain segregation with three C. parapsilosis isolates. The (GATA)4 probe was also used to detect restriction fragment length polymorphisms among a genetically closely related group of atypical Candida isolates on primary isolation from an additional HIV-infected patient. These chlamydospore-positive atypical Candida isolates were sucrose positive, were of C. albicans serotype A, hybridized weakly with the C. albicans-specific mid-repeat sequence probe 27A, and yielded fingerprint profiles by random polymorphic DNA analysis that were distinct from those derived from C. albicans isolates. The C. stellatoidea ex-type strain NCPF 3108 was indistinguishable from the atypical Candida isolates in all these tests and also yielded an identical carbohydrate and nitrogen source assimilation profile by using the ID 32C yeast identification system.

The abundance of various polymorphic microsatellite motifs differs between plants and vertebrates.

The abundance of different simple sequence motifs in plants was accessed through data base searches of DNA sequences and quantitative hybridization with synthetic dinucleotide repeats. Database searches indicated that microsatellites are five times less abundant in the genomes of plants than in mammals. The most common plant repeat motif was AA/TT followed by AT/TA and CT/GA. This group comprised about 75% of all microsatellites with a length of more than 6 repeats. The GT/CA motif being the most abundant dinucleotide repeat in mammals was found to be considerably less frequent in plants. To address the question if plant simple repeat sequences are variable as in mammals, (GT)n and (CT)n microsatellites were isolated from B.napus. Five loci were investigated by PCR-analysis and amplified products were obtained for all microsatellites from B. oleracea, B.napus and B.rapa DNA, but only for one primer pair from B.nigra. Polymorphism was detected for all microsatellites.

Physical and genetic characterization of the distal segment of the myotonic dystrophy area on 19q

The mutation involved in myotonic dystrophy (DM) has been mapped to the region between the ERCC1 DNA repair gene and the anonymous D19S51 locus on 19q13.3. Starting at locus D19S112 (probe pX75b), which served as a novel entry site for this chromosome region, we have established a cosmid contig of approximately 200 kb. In the contig, a gene expressed in the brain and a highly informative, 12-allele (TG) n variable simple sequence motif (VSSM) were identified. With this marker, designated X75b-VSSM, a highly characteristic size distribution of alleles linked with DM, which differed significantly from that on normal chromosomes, was observed. Combining our physical mapping and genetic data, we show that the X75b-VSSM marker is the closest distal to DM, thus excluding the DM mutation from the entire telomeric portion of the ERCC1-D19S51 region.

Ubiquitous nuclear factors bind specifically to a 5′-region conserved in carcinoembryonic antigen-related genes

We recently cloned members of the murine carcinoembryonic antigen (CEA) gene family, some of which are differentially expressed during placental development. By intra- and interspecies sequence comparisons, we identified an element in the putative promoter and/or 5′-nontranslated region which is conserved within all human and rodent CEA-related genes analyzed so far. Using gel retardation analysis and DNasel hypersensitive site mapping, we now show that ubiquitously expressed nuclear factors specifically bind to the conserved region derived from the mouse gene Cea-2 in vitro and probably also in vivo. Another DNasel hypersensitive site lies within or close to a simple sequence motif [(GGA) n] located in the first intron of Cea-2. Such sequences have been reported to play a role in the regulation of certain genes. Therefore, this analysis has identified putative regulatory regions for Cea-2 and possibly CEA-related genes in general.

Identification of variable simple sequence motifs in 19q13.2-qter: Markers for the myotonic dystrophy locus

Variable simple sequence motifs (VSSMs), or microsatellites, were used for the genetic delimitation of the myotonic dystrophy (DM) region at 19q. Three simple sequence motifs were identified in and around the ERCC1 DNA-repair gene at 19q13.2–13.3 and one in the vicinity of the RRAS gene at 19q13.3-qter. A (TG) n repeat, situated within the ninth intron of the ERCC1 gene, was converted into a highly informative multiallelic marker using PCR-mediated DNA amplification and high-resolution gel analysis. The structurally similar sequence motif in the RRAS gene yielded a marker system with only two alleles. Use of these VSSMs for linkage analysis and haplotyping in a selected set of DM families revealed that the DM gene is distal but close to the ERCC1 locus and can be excluded from the CKM-ERCC1 interval at 19q13.2. The order for RRAS and other distally located markers was established as DM-D19S50-[RRAS,KLK]-D19S22-ter.

Unexpected flexibility in an evolutionarily conserved protein-RNA interaction: genetic analysis of the Sm binding site.

Human autoantibodies of the Sm specificity recognize a conserved set of proteins found in the U class small nuclear ribonucleoproteins (U snRNPs), key trans-acting factors involved in the splicing of mRNA precursors. The Sm protein binding site in U snRNAs is unusual because of its single-stranded nature and its simple sequence motif (AU5-6GPu). Here we use genetics to probe this specific protein-RNA interaction by saturation mutagenesis of the Sm binding site of the Saccharomyces cerevisiae U5 snRNA. The assay system used to analyze these mutations takes advantage of a conditionally expressed U5 gene which does not support growth under non-permissive conditions; U5 genes containing Sm site mutations were tested for their ability to complement this lethal phenotype. Our results indicate that the Sm binding site is remarkably tolerant to mutation despite its high degree of conservation, suggesting that relatively few or redundant specific contacts can determine recognition of single-stranded RNA by protein. A complementary biochemical analysis of these mutants demonstrates that integrity of the Sm site is necessary for snRNP stability in vivo and in vitro.

The mechanism of antigen receptor gene assembly

Division of Biology California Institute of Technology Pasadena, California 91125 f Laboratory of Molecular Biology National Institute of Diabetes and Digestive and Kidney Diseases National Institutes of Health Bethesda, Maryland 20892 Much of the variety in binding specificities of the immune system is generated when subgenic DNA elements are assembled into antigen receptor genes during T and B cell differentiation. This process has attracted interest both because of its important role in the differentiation of the immune system and because it is the only site-specific recombination system known to operate in vertebrates. The natural substrates for immunoglobulin and T cell receptor gene rearrangement are complex loci (seven have been described in mice) composed of multiple cop- ies of V, D in some cases, and J segments. A complete an- tigen receptor gene is created by fusion of one V to one J, or one V to one D to one J. The basic operation (which we call V-J joining whether it involves joining V to J, V to D, or D to J), is represented in the left column of the figure. Two coding segments (shown as boxes) are detached from their joining signal sequences (shown as triangles) and then attached to one another to form a contiguous coding sequence. The junction is called a coding joint. A recipro- cal fusion of the signal sequences leads to a signal joint. (General principles are reviewed in Tonegawa, 1983.) The details of this process can be probed using recom- bination substrates that are constructed to contain a de- fined pair (or pairs) of gene segments. These substrates are designed either to become integrated into the genome of the recipient cells or to remain extrachromosomal. In ei- ther case, when introduced into recombinationally active cell lines, the gene segments are targeted by the recombi- nation machinery and become rearranged in the same fashion as their endogenous counterparts. The ease of experimentation afforded by this methodology has led to a rapid increase in available information about V-J recom- bination. Here we will review some of the key features of the process and present a current model of how it may work. One clearcut result is that the joining signals alone are the necessary and sufficient targets for recombination. Joining signals are simple DNA sequence motifs consist- ing of conserved heptamer and nonamer sequences separated by a nonconserved spacer of 12 or 23 bp. With- out joining signals, no recombination occurs. On the other hand, if V, D, or J coding elements are replaced by other DNA, the new sequences abutting the joining signals are still recombined site specifically (Lewis et al., 1985; Akira et al., 1987; Hesse et al., 1987). Thus, of the four elements

Use of variable simple sequence motifs as genetic markers: application to study of myotonic dystrophy

Among the many classes of repetitive elements present in the human genome, the ubiquitous "simple sequence motifs" (SSMs) composed of [A]n, [TG]n, [AG]n or codon-tandem repeats form a major source of genetic variation. Here we report a detailed molecular-genetic study of a "variable simple sequence motif" (VSSM) in the apolipoprotein C2 (apoC2) gene, which maps to the 19q13.2 region in the vicinity of the myotonic dystrophy (DM) locus. By combining in vitro DNA-amplification using the polymerase chain reaction and high-resolution gel electrophoresis, we could demonstrate a high degree of allelic variation with at least ten alleles, which differ in the number of repeated [TG] or [AG] dinucleotide units. Similar results were found for the somatostatin I gene locus. To evaluate the usefulness of SSM-length polymorphisms as genetic markers, the apoC2-VSSM was employed for linkage analysis in DM families. Our results establish that the orientation of the apolipoprotein gene cluster on 19q is cenapoE-apoC2-ter and indicate that the many thousands of structurally similar VSSMs in the human genome represent a rich source of highly informative genetic and diagnostic markers.

The Caenorhabditis elegans genome contains monomorphic minisatellites and simple sequences.

Many species have been shown to contain tandemly repeated short sequence DNA known as minisatellites and simple sequence motifs. Due to allelic variation in the copy number of the repeat unit these loci are usually highly polymorphic. Here we demonstrate the presence of sequences in the genome of the nematode Caenorhabditis elegans which are homologous to two sets of short sequence DNA. However, when two independent strains were compared no polymorphism for these sequences could be detected.

Molecular coevolution among cryptically simple expansion segments of eukaryotic 26S/28S rRNAs.

The set of "expansion segments" of any eukaryotic 26S/28S ribosomal RNA (rRNA) gene is responsible for the bulk of the difference in length between the prokaryotic 23S rRNA gene and the eukaryotic 26S/28S rRNA gene. The expansion segments are also responsible for interspecific fluctuations in length during eukaryotic evolution. They show a consistent bias in base composition in any species; for example, they are AT rich in Drosophila melanogaster and GC rich in vertebrate species. Dot-matrix comparisons of sets of expansion segments reveal high similarities between members of a set within any 28S rRNA gene of a species, in contrast to the little or spurious similarity that exists between sets of expansion segments from distantly related species. Similarities among members of a set of expansion segments within any 28S rRNA gene cannot be accounted for by their base-compositional bias alone. In contrast, no significant similarity exists within a set of "core" segments (regions between expansion segments) of any 28S rRNA gene, although core segments are conserved between species. The set of expansion segments of a 26S/28S gene is coevolving as a unit in each species, at the same time as the family of 28S rRNA genes, as a whole, is undergoing continual homogenization, making all sets of expansion segments from all ribosomal DNA (rDNA) arrays in a species similar in sequence. Analysis of DNA simplicity of 26S/28S rRNA genes shows a direct correlation between significantly high relative simplicity factors (RSFs) and sequence similarity among a set of expansion segments. A similar correlation exists between RSF values, overall rDNA lengths, and the lengths of individual expansion segments. Such correlations suggest that most length fluctuations reflect the gain and loss of simple sequence motifs by slippage-like mechanisms. We discuss the molecular coevolution of expansion segments, which takes place against a background of slippage-like and unequal crossing-over mechanisms of turnover that are responsible for the accumulation of interspecific differences in rDNA sequences.

Three transposed elements in the intron of a human VK immunoglobulin gene.

Two gene segments coding for the variable region of human immunoglobulin light chains of the kappa type (VK genes, ref. 2) were found to have unusual structures. The two genes which are called A6 and A22 are located in duplicated gene clusters. Their restriction maps are very similar. About 4 kb of the A22 gene region were sequenced. It turned out that the intron contains an insert with the characteristics of a transposed element. The inserted DNA of 1.2 kb length contains imperfect direct and inverted repeats at its ends; at the insertion site a duplication of five nucleotides was found. Within the inserted DNA one copy each of an Alu element and of the simple sequence motif (T-G)17 were identified. Also these two repetitive sequences are themselves flanked by short direct repeats. The major inserted DNA has no significant homology to published human nucleic acid sequences. The whole structure is interpreted best by assuming a sequential insertion of the three elements. The coding region of the VK gene itself has several mutations which by themselves would render it a pseudogene; we assume that the insertion event(s) occurred prior to the mutations. According to mapping and hybridization data A6 is very similar to A22.