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At least 1,279 records · Page 71Linked to original sources
PubMed1993

High throughput DNA sequencing using an automated electrophoresis analysis system and a novel sequence assembly program.

The combination of high-resolution scanning, image-databasing technology and sequence assembly software makes it possible to assemble contiguous overlapping sequences of DNA in a fraction of the time required by manual methods. This paper describes and evaluates an assembly program that rapidly generates contigs from scanned images and provides a unique ability to verify disagreements between overlapping strands or fragments of complementary DNA sequences on-screen, further increasing sequencing throughput.

Base Sequence↗
PubMed1996

Efficient preparation of short DNA sequence ladders potentially suitable for MALDI-TOF DNA sequencing.

Duplex probes with five-base single-stranded overhangs were developed for positional sequencing by hybridization [Broude et al., Proc Natl Acad Sci USA 91:3072-3076, 1994]. The partially duplex probes can be employed to capture single-stranded oligonucleotide targets and form primer-template complexes. Recently we showed that partially duplex probes can prime Sanger sequencing reactions on immobilized, but non-ligated long single-stranded targets (approximately 500 nucleotide) [Fu et al., Proc Natl Acad Sci, in press]. Here immobilized, non-ligated partially duplex probes were used to capture and sequence short single-stranded targets. This strategy is capable of rapidly preparing large numbers of samples for future mass spectrometric DNA sequencing.

Base Sequence↗
PubMed1996

A bovine mtDNA-binding protein to a conserved sequence adjacent to the termination associated sequence in the vertebrate mitochondrial displacement loop region.

We have found a sequence-specific mitochondrial (mt) DNA-binding protein in bovine mitochondria. The binding site designated Mt5 is adjacent to the termination associated sequence in the vertebrate mt displacement loop region and is interspecifically conserved. An UV-crosslinking and gel filtration demonstrated that a Mt5-binding protein (designated Mt5BP) exists in the mitochondria as a homodimer of an approximately 97-kDa polypeptide. The nucleotide sequence of Mt5 is homologous to that of the binding site for the recently reported mt pause-region binding protein (mtPBP) 1 from sea urchin embryo mitochondria. Gel mobility shift competition experiments indicated that Mt5BP can recognize human sequence couterparts for the mtPBP1-binding site. These results suggest that Mt5BP is related to mtPBP1 with respect to discontinuance of heavy-strand synthesis.

Animals↗
PubMed1999

Complete sequence determination of the mouse and human CTLA4 gene loci: cross-species DNA sequence similarity beyond exon borders.

CTLA4 (CD152), a receptor for the B7 costimulatory molecules (CD80 and CD86), is considered a fundamental regulator of T-cell activation. In this paper, we present the complete primary structure of the mouse and human CTLA4 gene loci. Sequence comparison between the mouse and the human CTLA4 gene loci revealed a high degree of sequence conservation both for homologous noncoding regions (65-78% identity) and for coding regions (72-98% identity), with an overall score of 71% over the entire length of the two genes. Of the CTLA4 genomic regions aligned, five simple repetitive elements were found in the mouse locus, whereas two simple repetitive sequences were localized on the human locus. RNA blot analysis of mouse and human primary tissues indicated that both CTLA4 and T-cell receptor transcripts were found in most organs with generally higher levels in lymphoid tissues. The conservation of CTLA4 gene patterning raises the possibility that constrained gene evolution of CTLA4 may be linked to conserved transcriptional control of this locus.

Abatacept↗
PubMed2004

Comparative sequence analysis of the Gdf6 locus reveals a duplicon-mediated chromosomal rearrangement in rodents and rapidly diverging coding and regulatory sequences.

Duplicated segments of genomic DNA can catalyze both gene evolution and chromosome evolution. Here we describe a rodent-specific duplication involving the Uqcrb gene, a cis-regulatory element for the Gdf6 gene, and a chromosomal rearrangement. Comparisons of Gdf6 sequences from several placental mammals and platypus revealed many strongly conserved regions flanking Gdf6 and the adjacent Uqcrb gene. However, in rat and mouse a synteny break resides approximately 70 kb upstream of Gdf6, such that Gdf6 and Uqcrb are on separate chromosomes. In rodents, Gdf6 and Uqcrb are both associated with homologous duplicons that may have catalyzed a rearrangement separating the two genes. However, the duplicon spanned both Uqcrb and a cis-regulatory element that controls Gdf6 transcription in limb skeletal joints. In mouse and rat, one duplicon now contains a degrading Uqcrb pseudogene but retains strongly conserved sequences within a Gdf6 enhancer. In contrast, the other duplicon has retained the intact Uqcrb gene and (in mouse) a copy of the Gdf6 enhancer that has acquired novel mutations. The duplicons have separately maintained distinct functions of the ancestral sequence, consistent with a "subfunction partitioning" evolutionary model. These findings also provide an example of a duplication that mobilized a tissue-specific enhancer from its cognate gene, and new evidence that duplications can be associated with chromosomal rearrangements. Furthermore, these data suggest that segmental duplications could lead to evolution of novel gene expression patterns via diversification of regulatory elements.

Animals↗
PubMed1996

Human skeletal muscle nebulin sequence encodes a blueprint for thin filament architecture. Sequence motifs and affinity profiles of tandem repeats and terminal SH3.

Analysis of deduced protein sequence and structural motifs of approximately 5500 residues of human fetal skeletal muscle nebulin reveals the design principles of this giant multifunctional protein in the sarcomere. The bulk of the sequence is constructed of approximately 150 tandem copies of approximately 35-residue modules that can be classified into seven types. The majority of these modules form 20 super-repeats, with each super-repeat containing a 7-module set (one of each type in the same order). These super-repeats are further divided into eight segments: with six segments containing adjacent, highly homologous super-repeats, one single repeat segment consisting of 8 nebulin modules of the same type, and a non-repeat segment terminating with a SH3 domain at the C terminus. The interactions of actin, tropomyosin, troponin, and calmodulin with nebulin fragments consisting of either repeating modules or the SH3 domain support its role as a giant actin-binding cofilament of the composite thin filament. Such affinity profiles also suggest that nebulin may bind to tropomyosin and troponin to form a composite calcium-linked regulatory complex on the thin filament. The modular construction, super-repeat structure, and segmental organization of nebulin sequence appear to encode thin filament length, periodicity, insertion, and sarcomere proportion in the resting muscle.

Actin Cytoskeleton↗
PubMed1999

Improved recognition of native-like protein structures using a combination of sequence-dependent and sequence-independent features of proteins.

We describe the development of a scoring function based on the decomposition P(structure/sequence) proportional to P(sequence/structure) *P(structure), which outperforms previous scoring functions in correctly identifying native-like protein structures in large ensembles of compact decoys. The first term captures sequence-dependent features of protein structures, such as the burial of hydrophobic residues in the core, the second term, universal sequence-independent features, such as the assembly of beta-strands into beta-sheets. The efficacies of a wide variety of sequence-dependent and sequence-independent features of protein structures for recognizing native-like structures were systematically evaluated using ensembles of approximately 30,000 compact conformations with fixed secondary structure for each of 17 small protein domains. The best results were obtained using a core scoring function with P(sequence/structure) parameterized similarly to our previous work (Simons et al., J Mol Biol 1997;268:209-225] and P(structure) focused on secondary structure packing preferences; while several additional features had some discriminatory power on their own, they did not provide any additional discriminatory power when combined with the core scoring function. Our results, on both the training set and the independent decoy set of Park and Levitt (J Mol Biol 1996;258:367-392), suggest that this scoring function should contribute to the prediction of tertiary structure from knowledge of sequence and secondary structure.

Models, Statistical↗
PubMed1993

Möbius sequence: further in vivo support for the subclavian artery supply disruption sequence.

Möbius sequence consists of a congenital bilateral facial nerve palsy and external ophthalmoplegia often associated with malformations of the limbs and orofacial structures. The pathogenesis of the sequence is a subject of debate. However, a new hypothesis proposes that Möbius sequence results from an interruption of embryonic blood supply (subclavian artery supply disruption sequence). Here we present an infant with bilateral facial nerve palsy (VII), external ophthalmoplegia (IV, VI), paresis of cranial nerves V, IX, X, XI, and XII, absence of the pectoralis major muscle (Poland anomaly), terminal transverse limb defects, and absence of the right diaphragm. Also, he was found to have discrete foci of brainstem calcifications in the region of the dorsal respiratory group on both CT scan and the histologic sections with microscopic evidence of diffuse brainstem "injury." The anomalies and histopathology noted in this infant imply that vascular insufficiency prior to the sixth week of gestation involving the proximal sixth intersegmental artery may result in the manifestations presented in this report and lend further support for the existence of a subclavian artery supply disruption sequence.

Abnormalities, Multiple↗
PubMed1983

Coding and 3' non-coding nucleotide sequence of chalcone synthase mRNA and assignment of amino acid sequence of the enzyme.

The nucleotide sequence of an almost complete cDNA copy of chalcone synthase mRNA from cultured parsley cells (Petroselinum hortense) has been determined. The cDNA copy comprised the complete coding sequence for chalcone synthase, a short A-rich stretch of the 5' non-coding region and the complete 3' non-coding region including a poly(A) tail. The amino acid sequence deduced from the nucleotide sequence of the cDNA is consistent with a partial N-terminal sequence analysis, the total amino acid composition, the cyanogen bromide cleavage pattern, and the apparent mol. wt. of the subunit of the purified enzyme.

Journal Article↗
PubMed1984

Complete nucleotide sequence of the infectious cloned DNA components of tomato golden mosaic virus: potential coding regions and regulatory sequences.

The nucleotide sequences of the infectious cloned DNA components of tomato golden mosaic virus (TGMV) have been determined. DNA A (2588 nucleotides) and DNA B (2508 nucleotides) have little sequence homology except for a region of 200 bases which is almost identical in the two molecules. Analysis of open reading frames revealed six potential coding regions for proteins of mol. wt. >10 000, four in DNA A and two in DNA B. Possible regulatory signals are identified and a model for bidirectional transcription of the two genome components is presented. Comparison of the nucleotide sequences of the DNAs of TGMV and cassava latent virus (CLV) revealed a fairly close relationship between TGMV DNA A and CLV DNA 1 and a comparatively distant relationship between TGMV DNA B and CLV DNA 2. All the potential coding regions in the TGMV DNAs had counterparts in the CLV DNAs suggesting an overall similarity in genome organisation, but six potential coding regions in the CLV DNAs had no counterparts in the TGMV DNAs. The 200-base region common to the two DNAs of each virus had little sequence homology, except for a highly conserved 33-base sequence potentially capable of forming a stable hairpin structure.

Journal Article↗
PubMed1995

Genetic recombination of pseudorabies virus: evidence that homologous recombination between insert sequences is less frequent than between autologous sequences.

We studied in vivo recombination between a thymidine kinase (TK) negative, glycoprotein E (gE) negative, attenuated strain and a virulent strain of pseudorabies virus (PRV) in pigs. To simplify the detection of recombination we inserted different but overlapping (375 bp) parts of the E1 gene of classical swine fever virus into the gG locus of both virus strains. Recombination between the E1 sequences of these viruses results in reconstitution of the complete E1 coding sequence and expression of the E1 protein. Since E1 is highly immunogenic, we expected to detect in vivo recombination in co-inoculated pigs by the presence of serum antibodies against E1. However, after co-inoculation of pigs with high doses of both virus strains, we were unable to detect antibodies against E1, suggesting that in vivo recombination did not occur or remained below the detection limit. Analysis of individual progeny viruses showed that 13 out of 995 (1.3%) possessed a recombinant TK-negative gE-positive phenotype. In contrast, no E1-positive viruses were detected among 5000 analyzed. This result showed that in vivo recombination between the two virus strains did occur, but was much more frequent between the TK and gE loci than between the E1 sequences. Similar results were obtained in in vitro recombination experiments in which possible growth differences between the various virus strains were excluded. The different recombination frequencies could not be attributed to the difference in distance of the genetic loci since recombination between mutations at a distance of 266 bp in the TK gene occurred as frequent as recombination between the TK and gE genes which are separated by approximately 60 kilobasepairs. These results indicate that some property of the E1 sequence and/or the location of the E1 sequence within the PRV genome affects the frequency of recombination.

Animals↗
PubMed2005

T1 Gd-enhanced compared with CISS sequences in retinoblastoma: superiority of T1 sequences in evaluation of tumour extension.

BACKGROUND: As adequate therapy for retinoblastoma in young children depends on infiltration of extra-retinal structures, diagnostic modalities play an essential role. METHODS: In this widely extended study, 80 children with retinoblastoma were studied with MRI (standard fat-suppressed Gd-enhanced T1, T2 thin-slice sequences (additionally with small loop surface coil), constructive interference in steady state (CISS) sequence covering the orbita). The images were analysed by two blinded neuroradiologists. Histology was used as the gold standard. RESULTS: MRI assumed infiltration of extra-retinal structures in 13 of 80 patients of which ten were confirmed by histology. Affected extra-retinal structures were: optic nerve (five, of which two were on CISS and three on T1 with higher image resolution using the surface coil), scleral infiltration (five, of which four on CISS and T1) and ciliary body infiltration (one on CISS and T1). Another 61 enucleated patients did not have any extra-retinal infiltration in histology. The CISS sequence with multiplanar reconstruction was mainly helpful in revealing exact three-dimensional tumour extension with excellent clinical acceptance and pre-surgical planning but T1 fat-suppressed Gd-enhanced images were superior in revealing exact tumour extension. CONCLUSION: CISS sequences allow to produce excellent anatomical images and to perform multiplanar reconstruction to better demonstrate tumour extension. However, T1-weighted sequences after contrast application are more sensitive (60 versus 40%) in detecting infiltration of the optic nerve but equal in detecting scleral infiltration.

Child↗
PubMed2003

Selective dendrite-targeting of mRNAs of NR1 splice variants without exon 5: identification of a cis-acting sequence and isolation of sequence-binding proteins.

Both protein and mRNA for the NR1 subunit of N-methyl-D-aspartate receptors are present in neuronal dendrites and undergo changes in distribution following synaptic excitation. However, the expression of all exonic splice variants of NR1 in dendrites has not been determined. In the present study, antibodies against the exon 5 (ex5) peptide sequence labeled proteins mostly in the soma of hippocampus neurons, whereas antibodies against ex21 or ex22 labeled cell bodies and dendrites. Antisense cRNAs for ex5 hybridized with mRNAs in cell bodies, whereas cRNAs for ex21 with mRNAs in both cell bodies and dendrites. Antisense DNA to a 24-base sequence identified as being present only in the 5'-UTR of cDNAs lacking ex5 (ex5(-)), hybridized with mRNAs in soma and dendrites and this labeling was coincident, mostly, with RNA granules. Insertion of the 24-base DNA ahead of that for enhanced green fluorescent protein (EGFP) increased the transport of EGFP mRNA and the expression of EGFP in neurites of neurons in culture. Fluorescent sense mRNA that contained the 24-base sequence bound to proteins in dendrites and to two proteins, 60 and 70 kDa, in brain microsomes. Proteins of similar size were also labeled by [32P]CTP-mRNA for NR1-(1a), which contains the 24-base 5'-UTR sequence, but not for NR1-(2b), which does not. Biotinylated 24-base sense mRNA was used to purify from brain microsomes two RNA-binding proteins (60 and 70 kDa). We concluded that the 24-base sequence in 5'-UTR of ex5(-) mRNA functioned as a cis-acting, dendrite-targeting element recognized selectively by two microsome proteins.

Animals↗
PubMed2001

Effect of sequence length, sequence frequency, and data acquisition rate on the performance of a Hadamard transform time-of-flight mass spectrometer.

Various factors influencing the performance of a Hadamard transform time-of-flight mass spectrometer (HT-TOFMS) have been investigated. Using a nitrogen corona discharge to produce an ion stream of N2+, N3+, and N4+, it is found for spectra containing only N4+ that the signal-to-noise ratio (SNR) closely approaches the value calculated from the ion background by assuming that the ion background follows a Poisson distribution. In contrast, for a more intense beam containing N2+, N3+, and N4+, the SNR is less than its theoretical value because of the appearance of discrete spikes in the mass spectrum caused by deviations in the actual modulation sequence from the ideal one. These spikes can be reduced, however, by decreasing the modulation voltage. Under these optimized conditions, the pseudo-random sequence length is varied to understand how it alters SNR, mass resolution, and scan speed. When the length of the pseudo-random sequence is doubled, the SNR increases by the square root of 2 while the time necessary to record a mass spectrum also doubles. Mass resolution can be varied between 500 and 1200 at m/z = 609 as the sequence length, modulation speed (10 MHz, 25 MHz), and acquisition rate (up to 50 MHz) are changed. Scan speeds of 6000 passes per s can be obtained using a sequence containing 4095 elements modulated at 25 MHz. The capability to tailor the HT-TOFMS to increase the scan speed and resolution with a constant 50% duty cycle makes the technique extremely appealing as a mass analyzer for measuring rapid changes in the composition of an ion stream.

Algorithms↗
PubMed2002

Identification and sequencing of HLA-B*0714 and B*2718 alleles and novel exon 1 sequences of B*0709 and B*2714 alleles in potential bone marrow donors.

Sequence specific oligonucleotide probe hybridization and sequence specific primer PCR typing of volunteer bone marrow donors suggested the presence of variants of known HLA-B alleles in two individuals. PCR products encompassing HLA-B locus exons 1, 2, and 3 were prepared, subcloned and sequenced. A Hispanic individual had a novel B*07 allele (B*0714) and a Chinese individual had a novel B*27 allele (B*2718). In two other individuals, a previously unknown sequence of exon 1 was determined for HLA-B*0709 (African American) and B*2714 (Native American). These findings further illustrate the substantial genetic variation present at the HLA-B locus within human populations. We discuss the structural variation in the protein sequence for these HLA-B alleles and its potential functional effects.

Asian People↗
PubMed1999

Perceptual-motor sequence learning of general regularities and specific sequences.

Participants viewed digit strings and typed them on a computer keyboard. When they used the same key configuration across training and test, they typed test strings that adhered to the same sequence rule as training strings faster than test strings that adhered to the opposite rule (general-regularity [GR] learning), and they typed test strings that were processed repeatedly during training faster than test strings that were not (specific-sequence [SS] learning). However, when they used different key configurations at training and at test, GR learning, but not SS learning, was observed. Conversely, when they did not type but spoke the strings aloud during training, SS learning, but not GR learning, was observed. Results suggest that in addition to declarative memory for specific sequences, relatively independent subsystems underlie procedural learning of perceptual-motor sequence components (producing GR effects) and sequence wholes (producing SS effects).

Female↗
PubMed2005

Targeted rearrangement of a chromosomal repeat sequence by transfection of a homologous DNA sequence using purified integrase.

Using a liposomal transfection with purified bovine leukemia virus (BLV) integrase, we observed an efficient DNA rearrangement of a chromosomal repeat sequence and targeted integration of a part of the transfected plasmid. The BLV integrase recognition sequence (IRS) including the 3' end of the BLV LTR U5, one of the sites cleaved by the integrase, was essential for the DNA rearrangement, and a sequence homologous to the chromosomal DNA neighboring the repeat target site had to be placed downstream of the IRS on the transfected plasmid. The pSV2neo DNA, including the pBR322 sequence preintegrated into L929 cells (primary transfectants), was rearranged by a secondary transfection of a pBR322-based hygromycin-resistance plasmid carrying the IRS. We present a model to explain the chromosomal DNA rearrangement of the primary clones through a homologous recombination-like reaction and amplification of the neighboring sequences.

Animals↗