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[Sexual reproduction of insects is regulated by cytoplasmic bacteria].

The effects have been considered that the intracellular symbiotic alpha-proteobacteria Wolbachia pipientis induces in its hosts, such as insects and other arthropods: cytoplasmic incompatibility upon mating, feminization, parthenogenesis, and androcide. Specific features of the bacterium genome and possible mechanisms of its action on hosts are discussed.

Animals↗

[Study on the molecular epidemiology of Mycobacterium tuberculosis in Shanghai].

OBJECTIVE: To explore the molecular-epidemiology of Mycobacterium tuberculosis in Shanghai. METHODS: Drug-resistant and drug-susceptible strains of M. tuberculosis were randomly selected from the bank of M. tuberculosis of Shanghai Municipal Center for Disease Control and Prevention and were genotyped by mycobacterial interspersed repetitive units(MIRU) and Spoligotyping methods. The genotyping results were analyzed and combined with epidemiological data. RESULTS: The Spoligotyping results demonstrated that 89 % (81/91) of the strains belonged to the Beijing genotype. Of the patients who had received BCG-vaccination,88.5% (54/61) infected with strains of Beijing genotype and 90.0% (27/30) of the patients were not BCG-vaccinated. However, the difference was not statistically significant. Drug-resistant rate from those strains of Beijing genotype was 45.7 (37/81), lower than that of non-Beijing genotype (60.0% ,6/10). Again,the difference was not statistically significant. The MIRU results showed that 62.6 % (57/91) were strains of clusters. CONCLUSION: The Beijing genotype of M. tuberculosis were found to be the dominant strains in Shanghai. The associations between Beijing genotype strains and BCG vaccination or drug-resistant were not found. Results from cluster analysis suggested that some cases might belong to the newly developed cases.

Adolescent↗

[Analysis of molecular profiles among Trypanozoon species and subspecies by MGE-PCR method].

OBJECTIVE: To analyze the relationship between genetic variability and evolution among Trypanosoma brucei (including T. b. brucei, T. b. rhodesiense and T. b. gambiense), T. evansi and T. equiperdum isolates. METHODS: Genomic DNAs of 26 trypanosome isolates were amplified by a mobile genetic elements (MGE) -PCR technique and cluster analysis was performed based on the molecular profiles with Neighbor-Joining method. RESULTS: The genetic variability among trypanosome isolates examined was obvious with an average genetic distance of 41.2% (ranged from 0 to 100%). Similarity coefficient among T. brucei isolates was 41.15% which was lower than that between T. evansi and T. equiperdum isolates. The closest relationship was found between T. evansi and T. brucei isolates with a similarity coefficient of 62.94%. The genetic variability between T. b. rhodesiense and T. b. brucei isolates was higher than that among T. b. gambiense isolates. CONCLUSION: Species and subspecies in Trypanozoon displayed a higher genetic variability; T. equiperdum isolates collected from China and from South America, and T. evansi isolates from China and from South America, should have a similar origin.

Animals↗

DNA sequence organization in the common Pacific starfish Pisaster ochraceous.

The sequence arrangement of the genomic DNA from the common sea star Pisaster ochraceous has been examined. Reassociation kinetics at DNA fragment lengths of 300 base pairs (bp) indicate the presence of at least three repetitive components in this DNA. The majority of these repetitive sequences are reiterated over the range from 10 to 100's. Approximately one-third of the nucleotides are found in repetitive sequences. Analysis of the reassociation kinetics of 3000-bp DNA fragments demonstrates the interspersion of repetitive and unique DNA sequences. The hyperchromicity of 3000-bp fragments reassociated to low Cot values (the product of moles of nucleotide per litre and time in seconds), and the size distribution of S1 nuclease resistant DNA duplex in these reassociation products, indicate a short-period interspersion pattern in the starfish genome. Repetitive segments (400 +/- 100 bp) are interspersed with longer unique DNA sequences. At a fragment length of 3000 bp the major fraction of the single-copy DNA is found in such an arrangement. In addition to short repetitive segments a substantial portion of the repetitive DNA nucleotides are found in segments excluded by Sepharose CL-2B (greater than or equal to 2000 bp). As much as one-quarter of the repetitive sequence nucleotides can be assigned to long segments.

Animals↗

DNA sequence organization in the alga Euglena gracilis.

The sequence organization of nuclear DNA in the single-celled alga Euglena gracilis has been studied by a combination of techniques: (1) the comparison of the reassociation kinetics of DNA fragments 300, 2000 and 8100 nucleotides long; (2) the reassociation of 32P-labeled DNA fragments of various lengths with driver fragments 300 nucleotides long; (3) the hyperchromicity of DNA structures formed by the reassociation of repetitive sequences; (4) and the direct measurement of the size of the duplex regions of reassociated repetitive DNA resistant to S1 nuclease. The single copy DNA sequences are approximately 1500 nucleotide pairs long and are interspersed with repetitive DNA sequences. The repetitive DNA, consisting of both highly repetitive and middle repetitive sequences, consists of one fraction of nucleotide sequences (0.67) with an average size of 4900 nucleotide pairs and a second fraction (0.33) with an average size of 1000 nucleotide pairs, 34% of the DNA consists of foldback sequences which are present on 45% of the DNA 4000 nucleotides long.

Base Sequence↗

BOX-PCR is an adequate tool for typing Aeromonas spp.

PCR-based methods of fingerprinting take advantage of the presence of repetitive sequences that are interspersed throughout the genome of diverse bacterial species. They include the repetitive extragenic palindromic (REP) sequence, the enterobacterial repetitive intergenic consensus sequence (ERIC) and the 154-bp BOX element. The combination of the three methods is used for fine discrimination of strains and is designated as rep-polymerase chain reaction (PCR). REP-PCR and ERIC-PCR have been shown to be useful for typing Aeromonas strains. To our knowledge, rep-PCR fingerprinting method using the BOXA1R primer has never been tested on aeromonads. In this study, the BOX-PCR fingerprinting technique was evaluated for the discrimination of strains of some Aeromonas species. All strains were typeable and the majority showed unique banding patterns. Four strains from culture collections were used to investigate the reproducibility of the method. According to our results, BOX-PCR fingerprinting is applicable for typing of Aeromonas strains and can be considered as a useful complementary tool for epidemiological studies of members of this genus.

Aeromonas↗

A short interspersed repetitive element provides a new 3' acceptor site for trans-splicing in certain ribosomal P2 beta protein genes of Trypanosoma cruzi.

Four Trypanosoma cruzi genomic DNA fragments carrying different TcP2 beta genes have been isolated and sequenced. Three of them had a single TcP2 beta gene, while the 3.8-kb-long DNA segment encoding the TcP2 beta-H1.8 locus showed two TcP2 beta genes arranged in tandem. These genes were physically connected by a 428-bp-long DNA sequence that was also located immediately 5' to the first gene and immediately 3' to the second. Comparison of the 4 TcP2 beta gene loci, suggested that the insertion of this repeated element originated the duplication of its target sequence, a poly(dT) stretch. Approximately 1200 copies of this short sequence, named short interspersed repetitive element (SIRE), were found scattered in the genome. Analysis of the 5' non-coding regions of different TcP2 beta mRNAs, and RNA-PCR experiments suggested that the insertion of a SIRE upstream of a TcP2 beta-H1.8 gene introduced a new 3' spliced leader (SL) acceptor site in the TcP2 beta-H1.8 pre-mRNAs, encoded within the SIRE. Consequently, in the mature H1.8 mRNA the SL sequence is followed by 38 bases directly transcribed from the SIRE. Structural and functional features of this repeated element reveal similarity to the short interspersed repetitive DNA sequences detected in the genomes of several microorganisms.

Animals↗

Integration site preferences of the Alu family and similar repetitive DNA sequences.

Numerous flanking nucleotide sequences from two primate interspersed repetitive DNA families have been aligned to determine the integration site preferences of each repetitive family. This analysis indicates that both the human Alu and galago Monomer families were preferentially inserted into short d(A+T)-rich regions. Moreover, both primate repeat families demonstrated an orientation specific integration with respect to dA-rich sequences within the flanking direct repeats. These observations suggest that a common mechanism exists for the insertion of many repetitive DNA families into new genomic sites. A modified mechanism for site-specific integration of primate repetitive DNA sequences is provided which requires insertion into dA-rich sequences in the genome. This model is consistent with the observed relationship between galago Type II subfamilies suggesting that they have arisen not by mere mutation but by independent integration events.

Animals↗

Characterization of estrogen receptor variant mRNAs from human breast cancers.

The cDNAs for variant estrogen receptor (ER) mRNAs previously identified in human breast cancer biopsy samples have been cloned and characterized. Some of these cDNAs are unique to a tumor sample (e.g. clones 24 and 5), while others are present in multiple breast tumor samples (e.g. clone 4). The 5' ends of the variant cDNAs are essentially identical to sequences present in exons 1, 2, and 3 of the normal ER mRNA. However, at points which mark either the exon 2/intron or exon 3/intron boundaries, the variant cDNA sequences diverge and are unrelated to the normal ER mRNA. The unique sequences of clones 24 and 5 are unknown, and the unique sequence of clone 4 is related to the long interspersed repetitive LINE-1 sequences. The variant mRNAs contain open reading frames which could encode proteins containing known functional domains of the normal ER but missing others. In particular, the hormone binding domain of the normal ER is always missing. Furthermore, some of the variant transcripts may encode other unique proteins. In transient expression assays the proteins encoded by the variant ER mRNAs are unable to activate transcription of an estrogen-responsive reporter gene; neither are they able to modulate the ability of normal ER proteins to activate transcription.

Amino Acid Sequence↗

Rat LINE1: the origin and evolution of a family of long interspersed middle repetitive DNA elements.

We present approximately 7.0 kb of composite DNA sequence of a long interspersed middle repetitive element (LINE1) present in high copy number in the rat genome. The family of these repeats, which includes transcribing members, is the rat homologue of the mouse MIF-Bam-R and human Kpn I LINEs. Sequence alignments between specimens from these three species define the length of a putative unidentified open reading frame, and document extensive recombination events that, in conjunction with retroposition, have generated this large family of pseudogenes and pseudogene fragments. Comparative mapping of truncated elements indicates that a specific endonucleolytic activity might be involved in illegitimate (nonhomologous) recombination events. Sequence divergence analyses provide insights into the origin and molecular evolution of these elements.

Amino Acid Sequence↗

Two-dimensional gel electrophoretic analysis of the HindIII 1.8-kb repetitive-sequence family in the human genome.

The structure and organization of a human repetitive DNA family containing the HindIII 1.8-kb repetitive sequence were studied, using two-dimensional (2D) gel electrophoresis. The HindIII 1.8-kb sequence proved to be part of a repetitive sequence about 5 kb long and interspersed on the genome. The long repetitive sequence family contained several subgroups, as based on polymorphism of the restriction site. Recombinant phages containing the long repetitive sequence were isolated from the human genomic DNA library. Heteroduplex and restriction analysis showed that the structure of the repetitive sequence carried by the phages was close to that expected from 2D gel electrophoretic analysis. The 2D gel electrophoretic analysis was shown to be a reliable and useful approach for surveying and mass analysis of repetitive sequence families.

DNA Restriction Enzymes↗

Identification of novel simple sequence length polymorphisms (SSLPs) in mouse by interspersed repetitive element (IRE)-PCR.

Interspersed repetitive element (IRE)-PCR is a useful method for identification of novel human or mouse sequence tagged sites (STSs) from contigs of genomic clones. We describe the use of IRE-PCR with mouse B1 repetitive element primers to generate novel, PCR amplifiable, simple sequence length polymorphisms (SSLPs) from yeast artificial chromosome (YAC) clones containing regions of mouse chromosomes 13 and 14. Forty-two IRE-PCR products were cloned and sequenced from eight YACs. Of these, 29 clones contained multiple simple sequence repeat units. PCR analysis with primers derived from unique sequences flanking the simple sequence repeat units in seven clones showed all to be polymorphic between various mouse strains. This novel approach to SSLP identification represents an efficient method for saturating a genomic interval with polymorphic genetic markers that may expedite the positional cloning of genes for traits and diseases.

Animals↗

A cloned AluI-equivalent repetitive sequence that detects transfected hamster DNA.

An AluI-equivalent interspersed repetitive element showing 75% homology to a published consensus sequence, which is present at a level of 3 x 10(5) copies per hamster genome [Haynes et al., Mol. Cell. Biol. 1 (1981) 573-583], has been cloned into plasmid pUC8 as a tandem copy which can be readily excised, self-ligated and labelled for use as a hybridisation probe. This probe successfully detects integrated hamster sequences in the DNA of transfected human cells [Arrand et al., Proc. Natl. Acad. Sci. USA 86 (1989) 6997-7001], whereas repetitive hamster DNA isolated as a C0t = 1 (fast reannealing) fraction fails to do so. Hamster DNA-containing clones in genomic libraries derived from hamster/human hybrids are also easily detected with this probe, thus facilitating the isolation of hamster genes which complement human gene defects.

Animals↗

Repetitive sequence environment distinguishes housekeeping genes.

Housekeeping genes are expressed across a wide variety of tissues. Since repetitive sequences have been reported to influence the expression of individual genes, we employed a novel approach to determine whether housekeeping genes can be distinguished from tissue-specific genes by their repetitive sequence context. We show that Alu elements are more highly concentrated around housekeeping genes while various longer (>400-bp) repetitive sequences ("repeats"), including Long Interspersed Nuclear Element-1 (LINE-1) elements, are excluded from these regions. We further show that isochore membership does not distinguish housekeeping genes from tissue-specific genes and that repetitive sequence environment distinguishes housekeeping genes from tissue-specific genes in every isochore. The distinct repetitive sequence environment, in combination with other previously published sequence properties of housekeeping genes, was used to develop a method of predicting housekeeping genes on the basis of DNA sequence alone. Using expression across tissue types as a measure of success, we demonstrate that repetitive sequence environment is by far the most important sequence feature identified to date for distinguishing housekeeping genes.

3' Flanking Region↗

Genome comparison in the genus Mus: a study with B1, MIF (mouse interspersed fragment), centromeric, and Y-chromosomal repetitive sequences.

Using four repetitive sequences, we compared DNAs isolated from Mus caroli, M. cookii, M. hortulanus, M. musculus, M. pahari, M. saxicola, and M. spretus. Except for B1, these probes showed species-specific hybridization patterns. Mouse interspersed fragment (MIF) sequences were present in all species examined, but those defined by the 1.3-kb EcoR1 band were fewer in M. pahari and M. saxicola than in the other species. The Y-chromosomal probe showed male-specific accumulation only in M. hortulanus, M. musculus, and M. spretus, which are known to be closely related. The genetic difference between M. spretus and the other two species (M. hortulanus and M. musculus) was clearly demonstrated by a M. musuclus centromeric sequence that hybridized strongly to M. hortulanus and M. musculus DNA but was underrepresented in the genome of M. spretus. These results may suggest the usefulness of these repetitive sequences in the classification of Mus species that display only subtle morphological differences.

Animals↗

Sequence and expression of grasshopper antennapedia: comparison to Drosophila.

We have cloned and characterized the Antennapedia (Antp) gene from the grasshopper Schistocerca americana. The Antennapedia protein contains seven blocks of sequence, including the homeodomain, that are conserved in the homologous proteins of other insects, interspersed with (usually repetitive) sequences unique to each species. There is no similarity between 1.8 kb of 3' untranslated sequence in grasshopper and Drosophila. We examined Antennapedia protein expression in grasshopper using an antibody raised against a grasshopper fusion protein and reexamined its expression in Drosophila using several different antibodies. Early patterns of expression in the two insects are quite different, reflecting differing modes of early development. However, by the germband stage, expression patterns are quite similar, with relatively uniform epithelial expression throughout the thoracic and abdominal segments which later retracts to the thorax. Expression is observed in muscle pioneers, the peripheral nervous system, and the central nervous system (CNS). In the CNS expression is initially limited to a few neurons, but eventually becomes widespread. Both insects show strong expression in certain homologous identified neurons and similar temporal modulation of expression.

Amino Acid Sequence↗

Insertion of a short repetitive sequence (D88I) in a sea urchin gene: a typical interspersed repeat?

A comparison has been made between the Sp88 gene regions of the DNAs of the sea urchins Strongylocentrotus purpuratus (Sp.) and Strongylocentrotus drobachiensis (Sd.). Examination of the 3' terminal part of the transcribed region revealed a short repetitive sequence present in Sd. but absent from Sp. A 12-nucleotide sequence present once in Sp. is almost perfectly duplicated at both ends of the repeat in Sd., suggesting that a mobile repeat was inserted in the Sd. genome. Other members of this family of repeated sequences occur in many interspersed locations in the genomes of both species. Except for the insertion duplication, the inserted sequence lacks direct or reverse repeats.

Animals↗