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Results for “genomic tandem array”

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PubMed2006

TCP34, a nuclear-encoded response regulator-like TPR protein of higher plant chloroplasts.

We describe the identification of a novel chloroplast protein, designated TCP34 (tetratricopeptide-containing chloroplast protein of 34 kDa) due to the presence of three tandemly arranged tetratricopeptide repeat (TPR) arrays. The presence of the genes encoding this protein only in the genomes of higher plants but not in photosynthetic cyanobacterial prokaryotes suggests that TCP34 evolved after the separation of the higher plant lineage. The in vitro translated precursor could be imported into intact spinach chloroplasts and the processed products showed stable association with thylakoid membranes. Using a specific polyclonal antiserum raised against TCP34, three protein variants were detected. Two forms, T(1) and T(2), were associated with the thylakoid membranes and one, S(1), was found released in the stroma. TCP34 protein was not present in etioplasts and appeared only in developing chloroplasts. The ratio of membrane-bound and soluble forms was maximal at the onset of photosynthesis. The high molecular mass thylakoid TCP34 variant was found in association with a transcriptionally active protein/DNA complex (TAC) from chloroplasts and recombinant TCP34 showed specific binding to Spinacia oleracea chloroplast DNA. Two TCP34 forms, T(1) and S(1), were found to be phosphorylated. An as yet unidentified phosphorelay signal may modulate its capability for plastid DNA binding through the phosphorylation state of the putative response regulator-like domain. Based on the structural properties and biochemical analyses, we discuss the putative regulatory function of TCP34 in plastid gene expression.

Amino Acid Sequence↗

PubMed1996

GFP tagging of budding yeast chromosomes reveals that protein-protein interactions can mediate sister chromatid cohesion.

BACKGROUND: Precise control of sister chromatid separation is essential for the accurate transmission of genetic information. Sister chromatids must remain linked to each other from the time of DNA replication until the onset of chromosome segregation, when the linkage must be promptly dissolved. Recent studies suggest that the machinery that is responsible for the destruction of mitotic cyclins also degrades proteins that play a role in maintaining sister chromatid linkage, and that this machinery is regulated by the spindle-assembly checkpoint. Studies on these problems in budding yeast are hampered by the inability to resolve its chromosomes by light or electron microscopy. RESULTS: We have developed a novel method for visualizing specific DNA sequences in fixed and living budding yeast cells. A tandem array of 256 copies of the Lac operator is integrated at the desired site in the genome and detected by the binding of a green fluorescent protein (GFP)-Lac repressor fusion expressed from the HIS3 promoter. Using this method, we show that sister chromatid segregation precedes the destruction of cyclin B. In mad or bub cells, which lack the spindle-assembly checkpoint, sister chromatid separation can occur in the absence of microtubules. The expression of a tetramerizing form of the GFP-Lac repressor, which can bind Lac operators on two different DNA molecules, can hold sister chromatids together under conditions in which they would normally separate. CONCLUSIONS: We conclude that sister chromatid separation in budding yeast can occur in the absence of microtubule-dependent forces, and that protein complexes that can bind two different DNA molecules are capable of holding sister chromatids together.

Bacterial Proteins↗

PubMed2005

Genome-wide transcription analyses in rice using tiling microarrays.

Sequencing and computational annotation revealed several features, including high gene numbers, unusual composition of the predicted genes and a large number of genes lacking homology to known genes, that distinguish the rice (Oryza sativa) genome from that of other fully sequenced model species. We report here a full-genome transcription analysis of the indica rice subspecies using high-density oligonucleotide tiling microarrays. Our results provided expression data support for the existence of 35,970 (81.9%) annotated gene models and identified 5,464 unique transcribed intergenic regions that share similar compositional properties with the annotated exons and have significant homology to other plant proteins. Elucidating and mapping of all transcribed regions revealed an association between global transcription and cytological chromosome features, and an overall similarity of transcriptional activity between duplicated segments of the genome. Collectively, our results provide the first whole-genome transcription map useful for further understanding the rice genome.

Chromosomes↗

PubMedDate not supplied

Cloning of a major repeat DNA sequence from Pyura stolonifera.

We have cloned a 147-bp Hind III fragment from a marine ascidian Pyura stolonifera. This sequence is arranged in tandem in arrays up to 20 kb in size and represents more than 5% of the total genomic DNA. The basic 147-bp unit was isolated from Hind III-digested genomic DNA and cloned into M13. Sequence analysis of seven clones revealed that the sequence is AT rich (75%) and can be separated from main band DNA by equilibrium density gradient centrifugation in the presence of the ligand dye Hoechst 33258. The sequence is highly conserved and is changed only by single base substitution mutations in the different clones. Use of this sequence as a probe demonstrated varying degrees of hybridization with DNA isolated from a wide range of other ascidians. Northern blot analysis revealed the presence of RNA hybridizing to the repeat in unfertilized eggs but transcription of this sequence was not evident in the adult organism.

Animals↗

PubMed1986

The trypanosome spliced leader small RNA gene family: stage-specific modification of one of several similar dispersed genes.

Diverse mRNAs of Trypanosoma brucei possess the same 5' terminal 35 nucleotides, termed the spliced leader (SL), which appears to be derived from a separate 135 nucleotide transcript. This small SL RNA is encoded within a 1.4 kb unit of DNA which is tandemly reiterated in the genome. In addition, there are at least 4 orphon elements containing SL sequences dispersed from the tandem array. Here we show that during the trypanosome life cycle one of the SL orphons undergoes a stage-specific modification that prevents cleavage of an EcoRV site and we further demonstrate that although only one orphon is modified, three of the SL orphons are flanked by very similar sequences. Each of these contains SL reiteration units including the non-transcribed spacer DNA, suggesting that they did not originate through an RNA intermediate. In addition no evidence of direct repeats at the junction of 1.4 kb and non-1.4 kb DNA was observed. Finally, a phylogenetic survey indicates that while many trypanosomatid species possess similarly organized SL-like sequences, only the SL orphons of closely related subspecies of the T. brucei - T. evansi complex share similar flanking regions.

Animals↗

PubMed1990

A transcribed gene, containing a variable number of tandem repeats, codes for a human epithelial tumor antigen. cDNA cloning, expression of the transfected gene and over-expression in breast cancer tissue.

A monoclonal antibody, H23, that specifically recognizes a breast-tumor-associated antigen, was used to isolate a cDNA insert that codes for the antigenic epitope. Nucleotide sequencing of this cDNA, as well as a longer 850-bp cDNA insert, shows that they are composed of 60-bp (G + C)-rich tandem repeating units. The coding strand was determined and codes for a proline-rich 20-amino-acid repeat motif. A comparison of the highly conserved repeat unit with the deduced flanking amino acid sequences demonstrates conservation of specific subregions of the repeat consensus within the flanking amino acids. Hybridization of the 60-bp cDNA probe with RNAs extracted from a variety of primary and metastatic human tumors yields relatively high levels of hybrid with the breast carcinomas, as compared to lower hybrid levels with RNAs from other epithelial tumors. RNA extracted from breast tissue adjacent to the tumor or from benign breast tumors, demonstrates low or undetectable levels of hybridization. Probing Southern blots with the 60-bp repeat shows that the tumor antigen is highly polymorphic and contains a variable number of tandem repeats (VNTRs). The VNTR nature of the gene was confirmed by probing Southern blots with unique genomic sequences that are physically linked to an isolated gene fragment that also contains the tandem repeat array. Mouse cells transfected with this gene fragment produce tumor antigen that is readily detected by H23 monoclonal antibodies. The allelic forms seen in 10 different primary human tumors demonstrate 100% concordance with the various mRNA species expressed. These studies are extended to the protein forms detected by immunoblot analyses that show both a correlation of the expressed tumor antigen species with the allelic forms as well as significantly increased expression in breast cancer tissue. The above studies unequivocally establish the over-expression of a VNTR gene coding for an epithelial tumor antigen in human breast cancer tissue.

Amino Acid Sequence↗

PubMed1995

Characterization of a family of tandemly repetitive DNA sequences from the fern sawfly, Strongylogaster osmundae (Hymenoptera: Tenthredinidae).

A family of repetitive DNA sequences in the genome of the sawfly, Strongylogaster osmundae (Hymenoptera: Tenthredinidae) was characterized. The family consists of a tandemly arranged array whose basic repeat unit is 1.0 kb. According to the compositions and arrangements of bases, the basic repeat unit can be divided into three distinct domains. Three domains share nucleotide sequence homology of 88, 74 and 89%, respectively, between members in this family. Second domain which had lower homology with the other two domains was found characteristically rich with polypurine/polypyrimidine sequences.

Animals↗

PubMed1988

Molecular evolution of the mouse proline-rich protein multigene family. Insertion of a long interspersed repeated DNA element.

Proline-rich proteins (PRPs) in the salivary glands of mice, rats, and hamsters are encoded by tissue-specific inducible multigene families. Mouse PRP genes are located on chromosome 8, and transcription is dramatically induced (about 70-fold) by isoproterenol treatment. Clones containing two nonallelic PRP genes (MP2 and M14) were isolated from cosmid and phage libraries of CD-1 mouse genomic DNA. The cloned regions comprise a contiguous block of 77 kilobase pairs of the mouse genome. Restriction mapping established the physical lineage of PRP genes MP2 and M14, and they are tandemly arrayed. The DNA sequence analysis presented in this report suggests that genes M14 and MP2 (Ann, D. K., and Carlson, D. M. (1985) J. Biol. Chem. 260, 15863-15872) arose via a gene duplication of a common ancestor. Two major differences between M14 and MP2 were observed. PRP gene MP2 has 13 tandemly arrayed 42-nucleotide repeats in exon II, whereas M14 has 17 repeats, and PRP gene M14 has an insertion by transposition of a 2-kilobase pair member of the long interspersed repeated DNA (LINE) family (LIMd) into intron I. The evolution of this PRP multigene family has been dominated by intra-exonic amplification of repeating nucleotide units coding for these and other proline-rich repeated peptides and by gene duplication. The LIMd element gives rise to heterogenous EcoRI, BamHI, and HindIII restriction enzyme patterns, and this insertion is also present in BALB/c, C57BL/6J, and DBA/2J mice.

Amino Acid Sequence↗

PubMed1983

Genomic organization, DNA sequence, and expression of chicken embryonic histone genes.

We have isolated and characterized in detail 15 lambda Charon 4A recombinant bacteriophage containing histone genes from a chicken genomic library. Restriction enzyme-mapping analysis and Southern hybridization to sequenced, homologous histone probes indicate that these genes are not tandemly reiterated within the chicken genome; they usually reside in clusters even though there is no unique array of genes that appears to constitute a typical cluster. Chicken H4 and H1 genes were identified within the genomic recombinants and subsequently sequenced. Extensive regions of homology exist in the 5'- and 3'-flanking regions of the chicken H4 gene when compared to H4 genes from other organisms. In addition to the well documented histone-specific domains, two previously unreported regions of homology lie 5' to this gene: an octanucleotide and a pentanucleotide sequence lying 59 and 116 nucleotides upstream from the H4 gene CAP site, respectively. The H1 gene sequence predicts that the H1 polypeptide is 217 amino acids in length. The 5'-flanking domain of this gene contains, in addition to the transcriptional initiation site and the ATA box, two unusual sequences: one is a nonamer which resides 29 nucleotides upstream from the "ATA" box and is conserved in both the chicken and sea urchin H1 genes, while the other is a GC-rich repetitive sequence element. The majority of the chicken histone genes among the 15 unique lambda recombinant clones are expressed almost exclusively during in ovo development (i.e. from at least 4 days postfertilization up to hatching, about 20-21 days postfertilization) and appear not to be associated with any particular tissue type.

Amino Acid Sequence↗

PubMed2000

Tandemly repeated DNA is a target for the partial replacement of thymine by beta-D-glucosyl-hydroxymethyluracil in Trypanosoma brucei.

In the DNA of African trypanosomes a small fraction of thymine is replaced by the modified base beta-D-glucosyl-hydroxymethyluracil (J). The function of this large base is unknown. The presence of J in the silent variant surface glycoprotein gene expression sites and the lack of J in the transcribed expression site indicates that DNA modification might play a role in control of gene repression. However, the abundance of J in the long telomeric repeat tracts and in subtelomeric arrays of simple repeats suggests that J may also have specific functions in repetitive DNA. We have now analyzed chromosome-internal repetitive sequences in the genome of Trypanosoma brucei and found J in the minichromosomal 177-bp repeats, in the long arrays of 5S RNA gene repeats, and in the spliced-leader RNA gene repeats. No J was found in the rDNA locus or in dispersed repetitive transposon-like elements. Remarkably, the rDNA of T. brucei is not organized in long arrays of tandem repeats, as in many other eukaryotes. T. brucei contains only approximately 15-20 rDNA repeat units that are divided over six to seven chromosomes. Our results show that J is present in many tandemly repeated sequences, either at a telomere or chromosome internal. The presence of J might help to stabilize the long arrays of repeats in the genome.

Animals↗

PubMed1986

alpha DNA in African green monkey cells is organized into extremely long tandem arrays.

We have determined the size of arrays formed by tandemly repeated monomers of alpha DNA in African green monkey cells. DNA fragments containing intact alpha DNA arrays were generated by digestion of genomic DNA with restriction endonuclease that do not have sites in the alpha DNA consensus sequence. Their size was determined by Southern analysis and by sedimentation through neutral sucrose gradients followed by probing of each fraction for alpha sequences. The restriction fragments varied in size with the most frequent being 78 kilobase pairs long. We have also shown that they contain very little non-alpha DNA sequences. This suggests a minimum array of 450 tandemly repeated alpha DNA monomers, which is more than an order of magnitude larger than previously supposed.

Animals↗

PubMed1993

Isolation and identification of a novel tandemly repeated DNA sequence in the centromeric region of human chromosome 8.

EcoRI subclones, designated as 50E1 and 50E4, were independently obtained from a cosmid clone previously mapped to the centromeric region of human chromosome 8. Southern blot hybridization analyses suggested that both subclones contain repetitive DNA sequences different from the chromosome 8 specific alphoid DNA. DNA sequence analysis of the 704 bp insert of 50E1 and the 1,962 bp insert of 50E4 revealed that both inserts contained tandemly repeated units of approximately 220 bp. Fluorescence in situ hybridization studies confirmed these two subclones to be specifically located on the centromeric region of chromosome 8. A 220 bp consensus sequence, derived from nine monomeric repeats, showed no significant homology to alphoid consensus sequences or to other currently known human centromeric DNA sequence. Furthermore, no significant homology was found with any other DNA sequence deposited in the EMBL or GenBank databases, indicating that this chromosome 8 specific repetitive DNA sequence is novel. From slot blot experiments it was estimated that 0.013% of the human genome comprises 1,750 of these monomeric repeats, residing on the centromeric region of chromosome 8 in tandem array(s).

Base Sequence↗

PubMed2004

APOBEC3B and APOBEC3C are potent inhibitors of simian immunodeficiency virus replication.

In the human genome the apolipoprotein B mRNA-editing enzyme catalytic polypeptide (APOBEC)3 gene has expanded into a tandem array of genes termed APOBEC3A-G. Two members of this family, APOBEC3G and APOBEC3F, have been found to have potent activity against virion infectivity factor deficient (Deltavif) human immunodeficiency virus 1 (HIV-1). These enzymes become encapsidated in Deltavif HIV-1 virions and in the next round of infection deaminate the newly synthesized reverse transcripts. The lentiviral Vif protein prevents the deamination by inducing the degradation of APOBEC3G and APOBEC3F. We report here that two additional APOBEC3 family members, APOBEC3B and APOBEC3C, have potent antiviral activity against simian immuno-deficiency virus (SIV), but not HIV-1. Both enzymes were encapsidated in HIV-1 and SIV virions and were active against Deltavif SIV(mac) and SIV(agm). SIV Vif neutralized the antiviral activity of APOBEC3C, but not that of APOBEC3B. APOBEC3B induced abundant G --> A mutations in both wild-type and Deltavif SIV reverse transcripts. APOBEC3C induced substantially fewer mutations. APOBEC3F was found to be active against SIV and sensitive to SIV(mac) Vif. These findings raise the possibility that the different APOBEC3 family members function to neutralize specific lentiviruses.

Anti-HIV Agents↗

PubMed1989

Amplification of the ribonucleotide reductase small subunit gene: analysis of novel joints and the mechanism of gene duplication in vaccinia virus.

Amplification of the M2 gene encoding the small subunit of ribonucleotide reductase (EC 1.17.4.1) was analyzed in a collection of vaccinia virus (VV) isolates selected for resistance to 5 mM hydroxyurea (HU). Most of the mutants harbored tandem direct repeat arrays of the M2 gene, but several had duplicated M2 as an inverted repeat by genomic rearrangements involving the chromosomal termini. Novel joints formed by direct repeats were mapped, amplified in vitro, and sequenced. The junctions were simple fusions between DNA downstream and upstream of the M2 gene. Lack of sequence homology at the breakpoints indicated that the initial genomic rearrangements leading to gene amplification were due to nonhomologous recombination events.

Base Sequence↗

PubMedDate not supplied

[Genomic organization and chromosomal localization of a new repeat MS7, specific for heterochromatin of sex chromosomes in Microtus species voles of the arvalis group].

The tandemly arranged MS4 repeat with monomeric units of 4.1 kb is species-specifically distributed in heterochromatin of sex chromosomes of four common vole species of genus Microtus, group arvalis [1, 2]. In this work, we studied the genomic organization of the MS4 homolog in euchromatin of the X chromosome of M. arvalis. It has been shown by analyzing the phage genomic clones that one MS4 copy makes a part of a monomeric unit exceeding 8.5 kb that also includes a new MS7 repeat and, possibly, LINE fragments. MS7 is located together with MS4 in heterochromatin of common vole sex chromosomes, but in a substantially lesser amount. Probably, as a result of an evolutionary transition of an original repeat from euchromatin of the X chromosome to heterochromatin of the Y chromosome, MS4 underwent multiple amplification, and MS7 spread throughout heterochromatin, being surrounded by the MS4 tandem arrays.

Animals↗

PubMed1986

Evaluating quantitative variation in the genome of Zea mays.

Genomic diversity within the species Zea mays has been examined by measuring the variation in the repetitive component of the nuclear genome among North American inbred lines and varieties. This was done by preparing a set of clones of repetitive maize sequences that differ in function, molecular arrangement and multiplicity and then using these as probes for quantitative hybridization to DNA from various maize genotypes. The comparison showed that the majority of repeated sequences are markedly variable in copy number among the ten maize strains tested. The clone sample contained the rDNA and 5S genes, the major repeat of the chromosome knobs, sequences functioning as origins of DNA replication in yeast (ARS sequences) and randomly cloned sequences of unknown function and chromosomal location. The sequences ranged in reiteration frequency from 200 to greater than 10(5) copies and included both tandemly arrayed and dispersed repeats. The copy numbers were measured by hybridizing labeled cloned sequences to aliquots of high molecular weight genomic DNA that were applied to nitrocellulose filters through a slotted template (slot blotting). The hybridization signal on an autoradiogram occurred in a narrow band that could be scored reliably with a densitometer. This provided a rapid method of determining the abundance of particular repeated sequences in individual plants and plant populations. Using this technique, we found that the copy number of repeated sequences of all types generally varied among the strains by two- to threefold, although at least one sequence showed no detectable variation. In contrast to the variability found between strains, individuals within an inbred line or variety were found to be indistinguishable in terms of specific sequence multiplicity. Each genotype has a different pattern of copy numbers for the set of repeated sequence clones, and this pattern is characteristic of all individuals of a particular genotype. The data also show that the copy number of each sequence varies independently. No strains had uniformly high or low copy numbers for the entire set of probes.

DNA↗

PubMed2004

Formation and loss of large, unstable tandem arrays of the piggyBac transposable element in the yellow fever mosquito, Aedes aegypti.

The Class II transposable element, piggyBac, was used to transform the yellow fever mosquito, Aedes aegypti. In two transformed lines only 15-30% of progeny inherited the transgene, with these individuals displaying mosaic expression of the EGFP marker gene. Southern analyses, gene amplification of genomic DNA, and plasmid rescue experiments provided evidence that these lines contained a high copy number of piggyBac transformation constructs and that much of this DNA consisted of both donor and helper plasmids. A detailed analysis of one line showed that the majority of piggyBac sequences were unit-length donor or helper plasmids arranged in a large tandem array that could be lost en masse in a single generation. Despite the presence of a transposase source and many intact donor elements, no conservative (cut and paste) transposition of piggyBac was observed in these lines. These results reveal one possible outcome of uncontrolled and/or unexpected recombination in this mosquito, and support the conclusion that further investigation is necessary before transposable elements such as piggyBac can be used as genetic drive mechanisms to move pathogen-resistance genes into mosquito populations.

Aedes↗

PubMed1997

A family of highly repetitive DNAs from "ginbuna" (Carassius auratus langsdorfi) genome common to Carassius auratus populations.

A family of highly repetitive DNAs (Hi-a) in the genome from a population of the crucian carp, tentatively identified as the ginbuna (Carassius auratus langsdorfi), was isolated from the HindIII digests and characterized. The Hi-a monomer (268 or 269 bp) was AT-rich (64.9%) with internal repetitive oligomers. The nucleotide similarity among monomers within the same individual was 88-98%, whose sequence alterations occurred mostly at restricted sites. Hybridization analyses revealed that the Hi-a family was organized into tandem array(s) representing satellite DNAs, and that its variants that share certain restriction sites of the repeat unit were dispersed into the tandem array(s) of Hi-a DNAs at varying periodicities. The lack of sequences homologous to the ginbuna Hi-a in the genomes from cyprinid fishes other than gengorobuna and goldfish suggests that Hi-a DNAs are specifically present in the species C. auratus. The genome of goldfish differed somewhat in the quantity or the genomic organization of Hi-a DNAs from that of the ginbuna.

Animals↗