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PubMed1989

Construction of a Not I restriction map of the fission yeast Schizosaccharomyces pombe genome.

Pulsed field gel electrophoresis and large DNA technology were used to construct a Not I restriction map of the entire genome of the fission yeast Schizosaccharomyces pombe. There are 14 detectable Not I sites in S. pombe 972h: 9 sites on chromosome I and 5 sites on chromosome II, while no Not I sites were found on chromosome III. The 17 fragments (including intact chromosome III) generated by Not I digestion were resolved by PFG electrophoresis. These fragments ranged in size from 4.5 kb to approximately 3.5 Mb. Various strategies were applied in determining, efficiently, the order of the fragments on the chromosomes. The genomic size measured by adding all the fragments together is about 14 Mb and the sizes of the three chromosomes are I, 5.7 Mb, II, 4.6 to 4.7 Mb, and III, 3.5 Mb. These are generally somewhat smaller than estimated previously.

Blotting, Southern↗

PubMed1985

[Restriction map of the genetic transfer factor pAP42].

Based on the calculated molecular weights of EcoR1, HindIII, and SalI fragments of the genetic transfer factor pAP42 the restriction map of this plasmid was designed. Sites recognizing restrictases are mostly located in the plasmid fragment with a molecular weight of 5.7 MD.

Attachment Sites, Microbiological↗

PubMed1984

Restriction map of an agropine-type Ri plasmid and its homologies with Ti plasmids.

The Ri plasmid of an agropine-type Agrobacterium rhizogenes, strain HRI, was cloned in a cosmid and mapped with the restriction endonucleases BamHI, EcoRI, KpnI, SmaI, and XbaI. This plasmid is almost identical to pRi1855 and pRiA4b. A study by Southern hybridizations of the homologies with octopine pTiB6806 and nopaline pTiC58 makes it possible to propose the localization of certain functions on this plasmid, such as virulence, agropine catabolism, agropine synthesis, and the origin of replication.

Base Sequence↗

PubMed1991

Rapid restriction map constructions using a modified pWE15 cosmid vector and a robotic workstation.

This paper describes a number of techniques for rapid restriction mapping of cosmid clones. First, we have replaced the cloning site of cosmid vector pWE15 with a polylinker containing 15 infrequently cleaved restriction enzyme sites that are placed asymmetrically on each side of the BamHI cloning site. DNA cloned into this vector can be fully recovered by using several pairs of restriction enzymes. Second, we have designed a simple electrical circuit device that allows the performance of asymmetric voltage gradient field inversion gel electrophoresis (AFIGE) to improve the resolution of DNA molecules in the range of 20-50 kbp. AFIGE can be obtained by simply placing the device in between a commercially available switching unit and the gel box in a standard field inversion system. Finally, the restriction digestion procedure has been automated by using a Beckman Biomek 1000 robotic workstation. Using this automated system, 96 restriction reactions, including gel loading, can be performed in less than two hours. In summary, these methods represent at least a tenfold improvement in the speed and/or mapping data that can be obtained in a single gel.

Animals↗

PubMed1984

A restriction map of virulence plasmid pVYE439-80 from a serogroup 9 Yersinia enterocolitica strain.

A restriction map of the virulence plasmid pVYE439-80, isolated from Yersinia enterocolitica 439-80 (serogroup 9) was constructed for EcoRI, BamHI, SstII, and SmaI. The mapping was done after cloning of about two-thirds of the plasmid in Escherichia coli. The restriction pattern was compared to those obtained with plasmids isolated from Y. enterocolitica strains of serogroups 1, 3, and 5b. The restriction sites are particularly conserved in a region of about 25 kb. This region contains fragments that are also conserved in serogroup 8 strains [J. Heeseman, C. Keller, R. Morawa, N. Schmidt, H. J. Siemens, and R. Lauf (1983) J. Infect. Dis. 147, 107-115] and that were shown, in strains from this serogroup, to encode calcium dependency [D. A. Portnoy, H. Wolf-Watz, I. Bolin, A. B. Beeder, and S. Falkow, (1984) Infect. Immun. 43, 108-114].

Base Sequence↗

PubMed1992

Estimation of restriction maps with known site order using a generalized linear model.

A generalized linear model with Gamma errors is used to estimate the coordinates of a restriction map when the site order is known. This can be conveniently programmed in a wide range of statistical packages (e.g. Genstat 5, Minitab, SAS), and gives maximum likelihood estimates with their associated optimal properties. Regression diagnostics allow the checking of assumptions and help to identify mis-specified, influential or discordant fragment lengths. A specific diagnostic for identifying fragment lengths causing reversal of restriction site order is derived. Exact 'fragment' lengths from DNA sequencing can be conveniently included in an approximate manner by giving them a larger weight than observed restriction fragment lengths. Two examples and the Genstat 5 codes used in their analysis are presented.

Algorithms↗

PubMed1987

Restriction-map variation in natural populations of Drosophila melanogaster: white-locus region.

Restriction-map variation among 38 chromosomes collected from natural populations from around the world was surveyed using probes for a 45-kb region containing and surrounding the white locus. Insertion and deletion variation was more common in the regions flanking the white transcriptional unit, and restriction-site polymorphism appears to be most common 5' of the white locus. The frequencies of individual large insertions (suspected transposable elements) were low, although 37% of the chromosomes had at least one insertion in the white-locus region. The estimated level of nucleotide heterozygosity over the whole region was 0.012. There was little linkage disequilibrium among the polymorphic sites. In contrast to earlier reports of the variation in other regions of the Drosophila melanogaster genome, there seemed to be less linkage disequilibrium and perhaps more nucleotide polymorphism.

Animals↗

PubMed1987

Serotype-converting bacteriophage D3 of Pseudomonas aeruginosa: vegetative and prophage restriction maps.

D3 is a temperate serotype-converting bacteriophage of Pseudomonas aeruginosa. A restriction map, based upon BamHI, PstI, PvuI, HindIII and SmaI sites, indicates that the phage genome is 56.4 kb long, and that it possesses cohesive ends. The prophage map suggests a unique insertion site in the strain AK1380 genome. Phage DNA integration occurs upon the circularization of D3 genome with the integration point approximately equidistant from the two ends.

Bacteriophages↗

PubMed1988

Two human papillomavirus DNAs molecularly cloned from a patient with epidermodysplasia verruciformis: restriction maps.

Two distinct human papillomavirus (HPV) DNAs (MY-1 and MY-2) were molecularly cloned from the benign skin lesions of a patient with epidermodysplasia verruciformis. The restriction map of MY-1 was the same as that of HPV 3a. The map of MY-2 appeared to be different from those of any HPVs reported in the literature. MY-2 did not cross-hybridize with MY-1 or the DNAs of HPV types 1, 2 and 4 under stringent conditions.

DNA, Viral↗

PubMed1990

Human chromosome 17 NotI linking clones and their use in long-range restriction mapping of the Miller-Dieker chromosome region (MDCR) in 17p13.3.

A NotI linking library constructed from flow-sorted human chromosome 17 material was screened to aid in construction of a long-range restriction map of the Miller-Dieker chromosome region (MDCR) in 17p13.3. A total of 66 clones were mapped to one of eight regions of chromosome 17 using a somatic cell hybrid panel, and 44/66 (67%) of these clones cross-hybridized to rodent DNA on Southern blots. Of these, 24 clones were tested and all mapped to mouse chromosome 11, the homolog of human chromosome 17. Four linking clones mapped to 17p13.3 and were used for pulsed-field gel electrophoresis studies along with six other anonymous probes previously mapped to this region. Clone L132 was found to be deleted in all Miller-Dieker patients tested (n = 15) and therefore lies within the critical region for this disorder. It detects two NotI fragments (180 and 320 kb), one of which (320 kb) was shared by YNZ22 and YNH37, two probes previously shown to be co-deleted in all patients with the Miller-Dieker syndrome (MDS). These results indicate that all MDS patients share a minimum deletion region of greater than 370 kb. Two other NotI clones, L53 and L125, mapped telomeric to the MDS critical region and share a 600-kb MluI fragment with each other and with YNZ22/YNH37. This provides a 930-kb MluI map that encompasses the distal boundary of the MDS critical region but does not include the proximal boundary. A total of over 2 Mbp is represented in the MluI fragments by probes in subband p13.3, a cytogenetic region estimated to be 3-4 Mbp.

Abnormalities, Multiple↗

PubMed1988

Errors between sites in restriction site mapping.

Restriction site mapping programs construct maps by generating permutations of fragments and checking for consistency. Unfortunately many consistent maps often are obtained within the experimental error bounds, even though there is only one actual map. A particularly efficient algorithm is presented that aims to minimize error bounds between restriction sites. The method is generalized for linear and circular maps. The time complexity is derived and execution times are given for multiple enzymes and a range of error bounds.

Algorithms↗

PubMed1988

The rosy region of Drosophila melanogaster and Drosophila simulans. I. Contrasting levels of naturally occurring DNA restriction map variation and divergence.

A 40-kb region around the rosy and snake loci was analyzed for restriction map variation among 60 lines of Drosophila melanogaster and 30 lines of Drosophila simulans collected together at a single locality in Raleigh, North Carolina. DNA sequence variation in D. simulans was estimated to be 6.3 times greater than in D. melanogaster (heterozygosities per nucleotide of 1.9% vs. 0.3%). This result stands in marked contrast to results of studies of phenotypic variation including proteins (allozymes), morphology and chromosome arrangements which are generally less variable and less geographically differentiated in D. simulans. Intraspecific polymorphism is not distributed uniformly over the 40-kb region. The level of heterozygosity per nucleotide varies more than 12-fold across the region in D. simulans, being highest over the hsc2 gene. Similar, though less extreme, variation in heterozygosity is also observed in D. melanogaster. Average interspecific divergence (corrected for intraspecific polymorphism) averaged 3.8%. The pattern of interspecific divergence over the 40-kb region shows some disparities with the spatial distribution of intraspecific variation, but is generally consistent with selective neutrality predictions: the most polymorphic regions within species are generally the most divergent between species. Sequence-length polymorphism is observed for D. melanogaster to be at levels comparable to other gene regions in this species. In contrast, no sequence length variation was observed among D. simulans chromosomes (limit of resolution approximately 100 bp). These data indicate that transposable elements play at best a minor role in the generation of naturally occurring genetic variation in D. simulans compared to D. melanogaster. We hypothesize that differences in species effective population size are the major determinant of the contrasting levels and patterns of DNA sequence and insertion/deletion variation that we report here and the patterns of allozyme and morphological variation and differentiation reported by other workers for these two species.

Animals↗

PubMed1989

[Restriction map of E. coli shuttle plasmid (p# GTE5) with secretive function].

The shuttle plasmid (p# GTE5) DNA with secretive function was extracted by the alkali lysozyme method from E. coli RRI strain. Its molecular weight is 4.5 Md and DNA size is 6.9 Kb. Restriction fragments of plasmid was obtained by single and double enzymes complete digestion using five different restriction endonucleases. The restriction map of shuttle plasmid (p# GTE5) was established for the enzymes EcoRI, BglII, pstI, PvuII, and TaqI.

Escherichia coli↗

PubMed1987

Human adenovirus from subgenus D: restriction mapping of types 9 and 19 and characterization of a new genome type.

A strain of adenovirus (AV) was isolated from the stools of a child with acute gastroenteritis. This virus (strain 208) was specifically neutralized by antiserum to AV19, a species belonging to subgenus D and previously reported to cause keratoconjunctivitis. However, restriction analysis of strain 208 showed it to be distinct from AV19 and the other 40 human AV serotypes presently known. When calculated from 18 restriction patterns, the proportion of comigrating fragments common to AV19 and strain 208 was only 57%. This percentage was no higher than that obtained by comparison of strain 208 and AV9, a randomly chosen type from subgenus D. Strain 208 therefore appeared to represent a new genome type. The restriction maps of the three viruses for BamHI, ClaI, EcoRI, HindIII, MluI, NdeI and SfiI endonucleases are presented.

Adenoviruses, Human↗

PubMed1986

[Restriction mapping of recombinant plasmids carrying the genes for arginine biosynthesis in Escherichia coli K-12].

ArgA and argECBH genes of Escherichia coli K-12 were cloned on the pBR322 vector. Restriction maps of the recombinant plasmids were constructed. Deletion mutants of these recombinant plasmids, retaining the functional argA and argE genes, were obtained using different restriction enzymes. All of the recombinant derivatives have the replication properties of the pBR322 vector.

Arginine↗

PubMed1991

Long-range restriction mapping and linkage analysis of the Prader-Willi chromosome region (PWCR).

In an attempt to elucidate the relationship between genetic alterations at chromosomal bands 15q11.2-12 and the Prader-Willi syndrome (PWS), we have constructed a long-range restriction map of this region using a combination of pulsed-field gel techniques and the infrequently cutting restriction enzymes NotI, MluI, SalI, SfiI, NruI, SacII, and BssHII. Four previously reported probes mapping to 15q11.2-12 and known to be deleted in PWS patients were used to construct the physical map of this region. The loci recognized by these four probes have been localized to a 2600-kb partial SalI restriction fragment and a 3200-kb partial EcoRI restriction fragment. Linkage studies were performed on nine families to estimate the recombination rates between these loci. The calculated lod scores did not indicate significant linkage between any of the four loci. The contrast between the physical distance and the observed recombination frequency suggests that these four loci are located in a recombinational "hot spot."

Blotting, Southern↗

PubMed1978

pBR322 restriction map derived from the DNA sequence: accurate DNA size markers up to 4361 nucleotide pairs long.

I have derived a complete restriction map of pBR322 from the total nucleotide sequence of the plasmid. Most of the restriction sites also have been demonstrated empirically. The exact sizes of all restriction fragments and the relative positions of the cuts are presented. These fragments can serve as accurate DNA size markers from small pieces up to the 4362 base pair length of pBR322. Inserts cloned in this vector may be characterized easily using this data.

Base Sequence↗

PubMed1990

A long-range restriction map of the interleukin-4 and interleukin-5 linkage group on chromosome 5.

The genes for two of the hematopoietic growth factors, interleukin-4 and interleukin-5, are located on a small segment of chromosome 5 (q23-31), which is frequently deleted in myeloid disorders. Using pulsed-field gel electrophoresis, we demonstrate physical linkage of these two genes and present a long-range restriction map of the locus. The two genes are closely linked (maximum separation, 310 kb) and appear to be separated by an HTF island. We were unable to physically link these genes to two other closely related hematopoietic growth factor genes, interleukin-3 and granulocyte/macrophage colony-stimulating factor, which also map to this region of the genome. The clustering of these and other growth-related genes suggests that a higher order of genetic organization exists in this region of the chromosome.

Chromosomes, Human, Pair 5↗