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PubMed1981

Cloning, restriction endonuclease mapping and post-transcriptional regulation of rpsA, the structural gene for ribosomal protein S1.

Transducing lambda phages have been isolated that carry segments of the Escherichia coli chromosome in the aspC region, 20.5 min on the E. coli map. One of these phages, lambda aspC2, carries rpsA, the structural gene for the ribosomal protein S1. A three kilobase fragment from this phage, cloned into either the plasmid pACYC184 or the plasmid pBR322, was found to express S1. In cells carrying the rpsA gene on the high copy number plasmid pBR322 the rate of rpsA mRNA synthesis was increased 40-fold, whereas the rate of protein S1 synthesis was doubled, in comparison with these rates in an rpsA haploid.

Cloning, Molecular↗
PubMed1986

Molecular population genetics of the alcohol dehydrogenase gene region of Drosophila melanogaster.

Variation in the DNA restriction map of a 13-kb region of chromosome II including the alcohol dehydrogenase structural gene (Adh) was examined in Drosophila melanogaster from natural populations. Detailed analysis of 48 D. melanogaster lines representing four eastern United States populations revealed extensive DNA sequence variation due to base substitutions, insertions and deletions. Cloning of this region from several lines allowed characterization of length variation as due to unique sequence insertions or deletions [nine sizes; 21-200 base pairs (bp)] or transposable element insertions (several sizes, 340 bp to 10.2 kb, representing four different elements). Despite this extensive variation in sequences flanking the Adh gene, only one length polymorphism is clearly associated with altered Adh expression (a copia element approximately 250 bp 5' to the distal transcript start site). Nonetheless, the frequency spectra of transposable elements within and between Drosophila species suggests they are slightly deleterious. Strong nonrandom associations are observed among Adh region sequence variants, ADH allozyme (Fast vs. Slow), ADH enzyme activity and the chromosome inversion ln(2L)t. Phylogenetic analysis of restriction map haplotypes suggest that the major twofold component of ADH activity variation (high vs. low, typical of Fast and Slow allozymes, respectively) is due to sequence variation tightly linked to and possibly distinct from that underlying the allozyme difference. The patterns of nucleotide and haplotype variation for Fast and Slow allozyme lines are consistent with the recent increase in frequency and spread of the Fast haplotype associated with high ADH activity. These data emphasize the important role of evolutionary history and strong nonrandom associations among tightly linked sequence variation as determinants of the patterns of variation observed in natural populations.

Alcohol Dehydrogenase↗
PubMed1984

A restriction enzyme cleavage map of the histidine utilization (hut) genes of Klebsiella aerogenes and deletions lacking regions of hut DNA.

The histidine utilization (hut) operons of Klebsiella aerogenes were cloned into pBR322. The hut genes are wholly contained on a 7.9 kilobase pair fragment bounded by HindIII restriction sites and expression of hut is independent of the orientation of the fragment with respect to pBR322. A restriction map locating the 27 cleavage sites within hut for the enzymes, HindIII, PvuII, SalI, BglII, KpnI, PstI, SmaI, AvaI, and BamHI was deduced. Several of the cleavage sites for the enzymes HaeIII and HinfI were also mapped. A set of deletion plasmids was isolated by removing various restriction fragments from the original plasmid. These deletions were characterized and were used to assist in mapping restriction sites. This physical characterization of hut DNA opens the way for genetic and molecular analysis of the regulation of hut gene expression in vitro as well as in vivo.

Base Composition↗
PubMed1988

Physical map and polyhedrin gene sequence of Lymantria dispar nuclear polyhedrosis virus.

Restriction maps of the 166.6-kb genome of Lymantria dispar multiply-enveloped nuclear polyhedrosis virus (LdMNPV clone g) were constructed for BamHI, BglII, EcoRI, EcoRV, HindIII and KpnI, using cosmid pVK102 and pBluescript vectors. Southern hybridizations indicated that the LdMNPV genome contains five dispersed regions of intragenomic sequence homology. The polyhedrin gene of LdMNPV was located within BglII-E and the sequence of the 735-nucleotide (nt) coding region and 678 nt of flanking DNA was determined. A conserved 14-nt sequence, associated with transcriptional start points in other polyhedrins, was identified at 44 to 57 nt upstream from the start codon. The deduced polyhedrin amino acid (aa) sequence showed a high degree of homology with a previously determined protein sequence for LdMNPV polyhedrin (89%) and with deduced amino acid sequences for three other MNPV polyhedrins (74%). Optimal alignment of the four sequences indicated that LdMNPV polyhedrin possesses a single aa insertion at residue 4 and a single aa deletion at residue 164.

Amino Acid Sequence↗
PubMed1993

Physical genome map of the unicellular cyanobacterium Synechococcus sp. strain PCC 7002.

A physical restriction map of the genome of the cyanobacterium Synechococcus sp. strain PCC 7002 was assembled from AscI, NotI, SalI, and SfiI digests of intact genomic DNA separated on a contour-clamped homogeneous electric field pulsed-field gel electrophoresis system. An average genome size of 2.7 x 10(6) bp was calculated from 21 NotI, 37 SalI, or 27 SfiI fragments obtained by the digestions. The genomic map was assembled by using three different strategies: linking clone analysis, pulsed-field fragment hybridization, and individual clone hybridization to singly and doubly restriction-digested large DNA fragments. The relative positions of 21 genes or operons were determined, and these data suggest that the gene order is not highly conserved between Synechococcus sp. strain PCC 7002 and Anabaena sp. strain PCC 7120.

Cyanobacteria↗
PubMed1995

YAC and cosmid contigs spanning the Batten disease (CLN3) region at 16p12.1-p11.2.

A yeast artificial chromosome (YAC) contig has been constructed in 16p12.1-p11.2 that encompasses three loci (D16S288, D16S299, and D16S298) closely linked to the gene causing Batten disease or juvenile-onset neuronal ceroid lipofuscinosis (CLN3). The physical map has been ordered using 42 sequence tagged sites. Four genes, interleukin-4 receptor (IL4R), phenol-preferring phenol sulfotransferase (STP), monoamine-preferring phenol sulfotransferase (STM), and sialophorin (SPN), have been mapped to the YAC contig. A partial genomic restriction map has been constructed to confirm the order and distances between D16S298, predicted to be the locus closest to CLN3. The overlapping genomic clones are a valuable resource for cloning the Batten gene (CLN3) and other genes in the region.

Antigens, CD↗
PubMed1995

A cladistic analysis of phenotypic associations with haplotypes inferred from restriction endonuclease mapping or DNA sequencing. V. Analysis of case/control sampling designs: Alzheimer's disease and the apoprotein E locus.

Present-day associations between haplotypes at a candidate locus and phenotypes exist when phenotypically important mutations occurred at some point during the evolution of the current array of genetic variation. A cladistic statistical design can be defined that focuses power by using the evolutionary history of the candidate DNA region. This paper shows how cladistic methodology is used for the analysis of case/control data, a common sampling design in genetic/disease association studies. A worked example is presented of the associations for sporadic early and late-onset forms of Alzheimer's disease with the 19q13.2 chromosomal region that includes the loci for apoproteins E, CI, and CII. This analysis confirms earlier reports of a strong association of the ApoE epsilon 4 allele with Alzheimer's disease but indicates that it is premature to consider this association causal, particularly for early onset cases. Associations were also found with the epsilon 2 allele, as previously reported, and with the 1 allele at the ApoCI locus. However, this analysis indicates that it is inappropriate both statistically and medically to use single markers as risk predictors when haplotype data are available, even when the mutation leading to the marker is identified as having a strong phenotypic association.

Age of Onset↗
PubMed1993

[Physical mapping of human chromosomes with linking clones and jumping clones].

Physical maps facilitate the correlation of genetic linkage maps and actual genome structure, and contribute to the immediate access to disease genes in question. The introduction of pulsed-field gel electrophoresis (PFGE) made it possible to construct restriction maps in Mb order with rare-cutting enzymes. Linking clones and jumping clones are useful tools for constructing long-range and accurate maps. The linking clones are used to identify adjacent restriction fragments and the jumping clones are to identify neighboring restriction sites. The general strategy for physical mapping using these clones and our application to NotI restriction mapping of human chromosome 21 are described.

Animals↗
PubMedDate not supplied

Algorithms for optical mapping.

Optical mapping is a novel technique for determining the restriction sites on a DNA molecule by directly observing a number of partially digested copies of the molecule under a light microscope. The problem is complicated by uncertainty as to the orientation of the molecules and by erroneous detection of cuts. In this paper we study the problem of constructing a restriction map based on optical mapping data. We give several variants of a polynomial reconstruction algorithm, as well as an algorithm that is exponential in the number of cut sites, and hence is appropriate only for small number of cut sites. We give a simple probabilistic model for data generation and for the errors and prove probabilistic upper and lower bounds on the number of molecules needed by each algorithm in order to obtain a correct map, expressed as a function of the number of cut sites and the error parameters. To the best of our knowledge, this is the first probabilistic analysis of algorithms for the problem. We also provide experimental results confirming that our algorithms are highly effective on simulated data.

Algorithms↗
PubMed1988

Characterization of plasmid pAZ1 and the type III dihydrofolate reductase gene.

The plasmid pAZ1, which determines trimethoprim and sulfonamide resistance, was characterized by restriction endonuclease mapping. The restriction map was identical to that of the incQ plasmid RSF1010 over a 5.1-kbp region. The type III dihydrofolate reductase gene was cloned, and the DNA sequence was determined. The predicted protein had 162 amino acid residues, and it was more closely related to the gram-negative bacterial chromosomal dihydrofolate reductases than to other plasmid or vertebrate dihydrofolate reductases. Sequence identity was 51% with the Escherichia coli enzyme and 44% with the Neisseria gonorrhoeae enzyme.

Amino Acid Sequence↗
PubMed1999

An algorithm combining discrete and continuous methods for optical mapping.

Optical mapping is a novel technique for generating the restriction map of a DNA molecule by observing many single, partially digested, copies of it, using fluorescence microscopy. The real-life problem is complicated by numerous factors: false positive and false negative cut observations, inaccurate location measurements, unknown orientations and faulty molecules. We present an algorithm for solving the real-life problem. The algorithm combines continuous optimization and combinatorial algorithms, applied to a non-uniform discretization of the data. We present encouraging results on real experimental data.

Algorithms↗
PubMed2000

An algorithm combining discrete and continuous methods for optical mapping.

Optical mapping is a novel technique for generating the restriction map of a DNA molecule by observing many single, partially digested copies of it, using fluorescence microscopy. The real-life problem is complicated by numerous factors: false positive and false negative cut observations, inaccurate location measurements, unknown orientations, and faulty molecules. We present an algorithm for solving the real-life problem. The algorithm combines continuous optimization and combinatorial algorithms applied to a nonuniform discretization of the data. We present encouraging results on real experimental data and on simulated data.

Algorithms↗
PubMed1998

Characterization of a human homolog (BACH1) of the mouse Bach1 gene encoding a BTB-basic leucine zipper transcription factor and its mapping to chromosome 21q22.1.

Clinical interest in the genes on human chromosome 21, especially with respect to Down syndrome (DS), has provided a strong impetus for the creation of a transcript map of this chromosome. In an effort to identify new human genes on the basis of cDNA analysis, we found several cDNA clones that corresponded to chromosome 21-specific transcripts. One of these, ha2303, showed strong similarity to the murine transcription factor Bach1. We subsequently determined the entire nucleotide sequence of this cDNA clone and found it to contain the whole coding sequence. The gene, termed BACH1, encodes a 736-amino-acid polypeptide with 80.3% identity to the murine Bach1 protein and contains a Cap'n'collar (CNC)-type basic leucine zipper (bZip) domain and a protein interaction motif, the BTB domain. Northern blot analysis revealed that BACH1 was expressed in all tissues examined. Mapping using the NotI restriction map and the YAC contig map showed that the BACH1 gene is located at 21q22.1 between the NotI sites LA329 (D21S338) and LL60 (D21S389) and within approximately 400 kb of LA329. Both the prospective function and the chromosomal location suggest that this gene may be a DS candidate gene, contributing to certain DS phenotypes, and is possibly involved in certain features of monosomy 21.

Amino Acid Sequence↗
PubMed2002

Combined k-space q-space pulsed ESR imaging: mapping of restricted diffusion in (FA)(2)PF(6).

A (FA)(2)PF(6) crystal from the family of the quasi-one-dimensional organic conductors was selectively damaged by a beam of Helium ions with a slitted mask placed in the beam's trajectory. Pulsed ESR density weighted imaging of the damaged crystal revealed the appearance of regions where the ESR signal was absent. The one-dimensional motion of the charge carriers was thus restricted to the undamaged sections. The local charge carrier spin dynamics in these restricted areas was probed by combined k-space q-space pulsed ESR imaging. The local expected appearance of the restricted pulsed gradient spin echo (PGSE) "diffusive diffraction" effect is shown. The position of the diffraction minima is compatible with the density imaging results.

Journal Article↗
PubMed1981

Cloning of E. coli pnp gene from an episome.

Starting with an F' episome harboring a transposon inserted in the pnp gene (Portier 1980), we were able to identify an EcoRI restriction fragment carrying the pnp and argG genes. This fragment, from both wild-type and mutant episomes, was cloned ni pACYC184. The presence of argG on the fragment allowed positive selection of the desired clones in an auxotrophic strain (argG). A restriction map was established and a fragment of 3 megadaltons subcloned in the plasmid vector pBR322. The pnp gene corresponds to about 50% of this subcloned segment and was roughly located by deletion mapping. The direction of transcription and locations of the promotor and gene extremities were determined by analyzing proteins synthesized in "maxi-cells". In addition, the gene coding for a 10,000 dalton protein was found to reside adjacent to the beginning of the pnp structural gene. Strains carrying plasmids which express the pnp overproduce polynucleotide phosphorylase.

Arginine↗
PubMed1985

Frequency and types of deletional alpha+-thalassemia in northern Sardinia.

We determined by restriction mapping the frequency of the -alpha 3.7 determinant in a random sample of 48 adults in Northern Sardinia. We found a frequency of 0.18 +/- 0.04 and demonstrated that only type I crossover as determined by Apa I digestion (Higgs et al. 1984) is present. Moreover, we showed that this haplotype is not associated with an Rsa I polymorphism 5' to the alpha 2-globin gene. These data support the hypothesis of a unique origin of this deletion in Sardinia.

Adult↗