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1.
(TG)n uncovers a sex-specific hybridization pattern in cattle 总被引:2,自引:0,他引:2
Y Kashi F Iraqi Y Tikochinski B Ruzitsky A Nave J S Beckmann A Friedmann M Soller Y Gruenbaum 《Genomics》1990,7(1):31-36
Screening of a bovine genomic library with the human minisatellite 33.6 probe uncovered a family of clones that, when used to probe Southern blots of bovine genomic DNA digested with the restriction enzyme HaeIII or MboI, revealed sexually dimorphic, but otherwise virtually monomorphic, patterns among the larger DNA fragments to which they hybridized. Characterization of one of these clones revealed that it contains different minisatellite sequences. The sexual dimorphism hybridization pattern observed with this clone was found to be due to multiple copies of two tandemly interspersed repeats: the simple sequence (TG)n and a previously undescribed 29-bp sequence. Both repeats appear to share many genomic loci including autosomal loci. In contrast, Southern analysis of AluI- or HinfI-digested bovine DNA with the (TG)n repeat used as a probe yielded substantial polymorphism. These results show that (i) different minisatellites can be found in a cluster, (ii) both simple and more complex repeated sequences other than the simple quaternary (GATA)n repeat can be sexually dimorphic, and (iii) simple repeats can reveal substantial polymorphism. 相似文献
2.
Y Nakamura C Julier R Wolff T Holm P O''Connell M Leppert R White 《Nucleic acids research》1987,15(6):2537-2547
We have examined the structure and DNA sequence of a human genomic locus that consists of a large hypervariable region made up of repeats of a simple sequence. With several restriction enzymes, the locus shows many restriction fragments that vary quantitatively as well as qualitatively. Other restriction enzymes produce only a single, high-molecular-weight fragment at this locus. Almost all of the fragments are revealed with a simple sequence probe. Southern transfers of the high-molecular-weight restriction fragments produced by the restriction enzymes NotI and SfiI, resolved by pulsed-field gel electrophoresis, gave at most two fragments, demonstrated to be allelic, showing that the majority of the restriction fragments seen in the complex patterns are at a single locus. The estimated size of the region homologous to the probe varied from 250 to 500 kilobases. DNA sequencing indicated that the region consists of tandem repeats of a 40-base-pair sequence. Some homology was detected to the tandem repeating units of the insulin gene and the zetaglobin pseudogene hypervariable regions, and to the "minisatellite" DNA at the myoglobin locus. 相似文献
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B Boldyreff K Wehr R Hecht O G Issinger 《Biochemical and biophysical research communications》1992,186(2):723-730
Using the coding region of the human CK-2 alpha cDNA as a probe for screening a genomic mouse library, positive clones representing four different genomic loci were isolated. Partial DNA sequences of these loci encompassing the first 120 nucleotides of the putative coding region are reported. One positive clone was further analyzed by sequencing a 3.1 kb XbaI fragment. This clone displays the characteristics of a pseudogene, i.e. lack of introns and several nucleotide insertions and deletions. In its 3' region it contains a 91 bp large CT-rich stretch which consists of (CCTT) and (CT) repeats; in the 5' region three (CCCCCT) repeats. 相似文献
5.
We present evidence that a proportion of alleles at two human minisatellite loci is undetected by standard Southern blot hybridization. In each case the missing allele(s) can be identified after PCR amplification and correspond to tandem arrays too short to detect by hybridization. At one locus, there is only one undetected allele (population frequency 0.3), which contains just three repeat units. At the second locus, there are at least five undetected alleles (total population frequency 0.9) containing 60-120 repeats; they are not detected because these tandem repeats give very poor signals when used as a probe in standard Southern blot hybridization, and also cross-hybridize with other sequences in the genome. Under these circumstances only signals from the longest tandemly repeated alleles are detectable above the nonspecific background. The structures of these loci have been compared in human and primate DNA, and at one locus the short human allele containing three repeat units is shown to be an intermediate state in the expansion of a monomeric precursor allele in primates to high copy number in the longer human arrays. We discuss the implications of such loci for studies of human populations, minisatellite isolation by cloning, and the evolution of highly variable tandem arrays. 相似文献
6.
Martins C Wasko AP Oliveira C Porto-Foresti F Parise-Maltempi PP Wright JM Foresti F 《Cytogenetic and genome research》2002,98(1):78-85
In higher eukaryotes, the 5S ribosomal DNA (5S rDNA) is organized in tandem arrays with repeat units composed of a coding region and a non-transcribed spacer sequence (NTS). These tandem arrays can be found on either one or more chromosome pairs. 5S rDNA copies from the tilapia fish, Oreochromis niloticus, were cloned and the nucleotide sequences of the coding region and of the non-transcribed spacer were determined. Moreover, the genomic organization of the 5S rDNA tandem repeats was investigated by fluorescence IN SITU hybridization (FISH) and Southern blot hybridization. Two 5S rDNA classes, one consisting of 1.4-kb repeats and another one with 0.5-kb repeats were identified and designated 5S rDNA type I and type II, respectively. An inverted 5S rRNA gene and a 5S rRNA putative pseudogene were also identified inside the tandem repeats of 5S rDNA type I. FISH permitted the visualization of the 5S rRNA genes at three chromosome loci, one of them consisting of arrays of the 5S rDNA type I, and the two others corresponding to arrays of the 5S rDNA type II. The two classes of the 5S rDNA, the presence of pseudogenes, and the inverted genes observed in the O. niloticus genome might be a consequence of the intense dynamics of the evolution of these tandem repeat elements. 相似文献
7.
Structure of the human genomic glutathione S-transferase-pi gene 总被引:6,自引:0,他引:6
8.
Characterization and genetic mapping of simple repeat sequences in the tomato genome 总被引:18,自引:0,他引:18
Tomato genomic libraries were screened for the presence of simple sequence repeats (SSRs) with seventeen synthetic oligonucleotide probes, consisting of 2- to 5-basepair motifs repeated in tandem. GAn and GTn sequences were found to occur most frequently in the tomato genome (every 1.2 Mb), followed by ATTn and GCCn (every 1.4 Mb and 1.5 Mb, respectively). In contrast, only ATn and GAn microsatellites (n > 7) were found to be frequent in the GenBank database, suggesting that other motifs may be preferentially located away from genes. Polymorphism of microsatellites was measured by PCR amplification of individual loci or by Southern hybridization, using a set of ten tomato cultivars. Surprisingly, only two of the nine microsatellite clones surveyed (five GTn, three GAn and one ATTn), showed length variation among these accessions. Polymorphism was also very limited betweenLycopersicon esculentum andL. pennelli, two distant species. Southern analysis using the seventeen oligonucleotide probes identified GATAn and GAAAn as useful motifs for the detection of multiple polymorphic fragments among tomato cultivars. To determine the structure of microsatellite loci, a GAn probe was used for hybridization at low stringency on a small insert genomic library, and randomly selected clones were analyzed. GAn based motifs of increasing complexity were found, indicating that simple dinucleotide sequences may have evolved into larger tandem repeats such as minisatellites as a result of basepair substitution, replication slippage, and possibly unequal crossing-over. Finally, we genetically mapped loci corresponding to two amplified microsatellites, as well as nine large hypervariable fragments detected by Southern hybridization with a GATA8 probe. All loci are located around putative tomato centromeres. This may contribute to understanding of the structure of centromeric regions in tomato. 相似文献
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Genetic/genomic polymorphism, i.e. variations in DNA sequences are ideally assayed by direct nucleotide sequencing of a gene region or other homologous segment of the genome. An easier and cheaper approach, however, if the variants are analyzed by hybridization technology using restriction fragment length polymorphisms (RFLPs) or by detection of the number of tandem repeats (VNTR) of small DNA segments, the "minisatellites". In this study we describe results of the DNA analysis of repetitive sequences of human 6th chromosome by the application of a chemiluminescent labeled probes. The allele frequency distribution of polymorphic DNA sequences has been determined in unrelated individuals. The isolated genomic DNA was cut with Pst I restriction enzyme, size fractionated on agarose gel and hybridized with a chemiluminescent labeled D6 S132 probe. At this locus the Pst I cleaved DNA fragments are ranging from 1841 to 6098 base pairs (bp). Specific genetic pattern was characterized by more frequent fragments (3313 and 3884 bp), and the rarely occurring ones (clustered between 1841-2595 and 5227-6098 bp). Our study provides a further possibility for characterization of individual genomic patterns. 相似文献
11.
Molecular characterisation of a hypervariable region downstream of the human alpha-globin gene cluster. 总被引:32,自引:2,他引:32
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We have characterised an unusual, highly polymorphic region of DNA located 8-kb downstream of the human alpha-globin gene complex. This hypervariable region (alpha-globin 3' HVR) is composed of an array of 17-bp tandem repeats, the number of which differs considerably (70-450) from one allele to another. The sequence of the 17-bp repeats is highly conserved within and between alleles. Furthermore, this sequence identifies a core oligonucleotide [5'-GNGGGG(N)ACAG-3'] that is common to three previously characterised hypervariable regions. At reduced stringency, a probe to the 3' HVR detects a new family of multiallelic loci that will be of value in the study of human genetics. 相似文献
12.
Microsatellites: genomic distribution,putative functions and mutational mechanisms: a review 总被引:35,自引:0,他引:35
Microsatellites, or tandem simple sequence repeats (SSR), are abundant across genomes and show high levels of polymorphism. SSR genetic and evolutionary mechanisms remain controversial. Here we attempt to summarize the available data related to SSR distribution in coding and noncoding regions of genomes and SSR functional importance. Numerous lines of evidence demonstrate that SSR genomic distribution is nonrandom. Random expansions or contractions appear to be selected against for at least part of SSR loci, presumably because of their effect on chromatin organization, regulation of gene activity, recombination, DNA replication, cell cycle, mismatch repair system, etc. This review also discusses the role of two putative mutational mechanisms, replication slippage and recombination, and their interaction in SSR variation. 相似文献
13.
MOTIVATION: Complex genomes contain numerous repeated sequences, and genomic duplication is believed to be a main evolutionary mechanism to obtain new functions. Several tools are available for de novo repeat sequence identification, and many approaches exist for clustering homologous protein sequences. We present an efficient new approach to identify and cluster homologous DNA sequences with high accuracy at the level of whole genomes, excluding low-complexity repeats, tandem repeats and annotated interspersed repeats. We also determine the boundaries of each group member so that it closely represents a biological unit, e.g. a complete gene, or a partial gene coding a protein domain. RESULTS: We developed a program called HomologMiner to identify homologous groups applicable to genome sequences that have been properly marked for low-complexity repeats and annotated interspersed repeats. We applied it to the whole genomes of human (hg17), macaque (rheMac2) and mouse (mm8). Groups obtained include gene families (e.g. olfactory receptor gene family, zinc finger families), unannotated interspersed repeats and additional homologous groups that resulted from recent segmental duplications. Our program incorporates several new methods: a new abstract definition of consistent duplicate units, a new criterion to remove moderately frequent tandem repeats, and new algorithmic techniques. We also provide preliminary analysis of the output on the three genomes mentioned above, and show several applications including identifying boundaries of tandem gene clusters and novel interspersed repeat families. AVAILABILITY: All programs and datasets are downloadable from www.bx.psu.edu/miller_lab. 相似文献
14.
We describe a protocol that uses a bioinformatically optimized primer in an isothermal whole genome amplification (WGA) reaction. Overnight incubation at 37 degrees C efficiently generates several hundred- to several thousand-fold increases in input DNA. The amplified product retains reasonably faithful quantitative representation of unamplified whole genomic DNA (gDNA). We provide protocols for applying this isothermal primer extension WGA protocol in three different techniques of genomic analysis: comparative genomic hybridization (CGH), genotyping at simple tandem repeat (STR) loci and screening for single base mutations in a common monogenic disorder, beta-thalassemia. gDNA extracted from formalin-fixed paraffin-embedded (FFPE) tissues can also be amplified with this protocol. 相似文献
15.
The total number of microsatellite loci is considered to be at least 10-fold lower in avian species than in mammalian species. Therefore, efficient large-scale cloning of chicken microsatellites, as required for the construction of a high-resolution linkage map, is facilitated by the construction of libraries using an enrichment strategy. In this study, a plasmid library enriched for tandem repeats was constructed from chicken genomic DNA by hybridization selection. Using this technique the proportion of recombinant clones that cross-hybridized to probes containing simple tandem repeats was raised to 16%, compared with < 0·1% in a non-enriched library. Primers were designed from 121 different sequences. Polymerase chain reaction (PCR) analysis of two chicken reference pedigrees enabled 72 loci to be localized within the collaborative chicken genetic map, and at least 30 of the remaining loci have been shown to be informative in these or other crosses. 相似文献
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J Zonana J Gault K J Davies M Jones D Browne M Litt N Brockdorff S Rastan A Clarke N S Thomas 《American journal of human genetics》1993,52(1):78-84
X-linked hypohidrotic ectodermal dysplasia (EDA) has been localized to the Xq12-q13.1 region. A panel of genomic DNA samples from 80 unrelated males with EDA has been screened for deletions at seven genetic loci within the Xq12-13 region. A single individual was identified with a deletion at the DXS732 locus by hybridization with the mouse genomic probe pcos169E/4. This highly conserved DNA probe is from locus DXCrc169, which is tightly linked to the Ta locus, the putative mouse homologue of EDA. The proband had the classical phenotype of EDA, with no other phenotypic abnormalities, and a normal cytogenetic analysis. A human genomic DNA clone, homologous to pcos169E/4, was isolated from a human X-chromosome cosmid library. On hybridization with the cosmid, the proband was found to be only partially deleted at the DXS732 locus, with a unique junctional fragment identified in the proband and in three of his maternal relatives. This is the first determination of carrier status for EDA in females, by direct mutation analysis. Failure to detect deletion of the other loci tested in the proband suggests that the DXS732 locus is the closest known locus to the EDA gene. Since the DXS732 locus contains a highly conserved sequence, it must be considered to be a candidate locus for the EDA gene itself. 相似文献
18.
Genetic analysis of the hypervariable region flanking the human insulin gene. 总被引:4,自引:2,他引:2
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In order to study the mechanisms for the generation of length diversity within the 5' flanking region of the human insulin gene, we have isolated and sequenced a previously uncharacterized allele. This allele, of a size intermediate between those three already described in the literature, encompasses 1,156 base pairs (bp) and contains 81 reiterated tandem oligonucleotides of 14-15 bp each. Population analysis on 298 independently sampled individuals by Southern blotting of genomic DNA demonstrates that the polymorphic portion of the insulin 5' flanking region varies from 400 to more than 8,000 nucleotides, being encoded by from 30 to over 540 oligomeric repeats. Length variability 5' to the insulin gene is a result primarily of unequal crossing over, which generates an expansion or contraction in the number of tandem repeat units per chromosome. A similar mechanism probably accounts for nondispersed reiterated sequences at other loci in the human genome. 相似文献
19.
The repetitive sequence PisTR-A has an unusual organization in the pea (Pisum sativum) genome, being present both as short dispersed repeats as well as long arrays of tandemly arranged satellite DNA. Cloning, sequencing and FISH analysis of both PisTR-A variants revealed that the former occurs in the genome embedded within the sequence of Ty3/gypsy-like Ogre elements, whereas the latter forms homogenized arrays of satellite repeats at several genomic loci. The Ogre elements carry the PisTR-A sequences in their 3′ untranslated region (UTR) separating the gag-pol region from the 3′ LTR. This region was found to be highly variable among pea Ogre elements, and includes a number of other tandem repeats along with or instead of PisTR-A. Bioinformatic analysis of LTR-retrotransposons mined from available plant genomic sequence data revealed that the frequent occurrence of variable tandem repeats within 3′ UTRs is a typical feature of the Tat lineage of plant retrotransposons. Comparison of these repeats to known plant satellite sequences uncovered two other instances of satellites with sequence similarity to a Tat-like retrotransposon 3′ UTR regions. These observations suggest that some retrotransposons may significantly contribute to satellite DNA evolution by generating a library of short repeat arrays that can subsequently be dispersed through the genome and eventually further amplified and homogenized into novel satellite repeats. 相似文献
20.
Stringently controlled plasmids generate DNA fingerprint patterns in mammals when used at low hybridization temperatures. In order to develop a probe for use in paternity testing in cattle we screened a bovine, partial genomic plasmid library with the PCR-amplified ori region of plasmid P1. Of eight isolated clones one generated strong band patterns at high stringency in various mammalian species (data not shown). Sequence analysis revealed an imperfect, compound dinucleotide repeat region, which was PCR-amplified and cloned into the plasmid vector pUC19. Fingerprint results generated by this probe (termed pITZ1) in cattle are compared with the results generated by VNTR-probe pV47, which itself was developed by screening a human chromosome 16 library with tandem repeats of bacteriophage M13. Probe pITZ1 is useful in conjunction with other VNTR-probes for DNA fingerprinting in cattle and donkey populations. The dinucleotide repeat region responsible for the band patterns generated with pITZ1 is close to an Alu -like sequence, which may be involved in eukaryotic replication mechanisms. 相似文献