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PubMed1982

Restriction enzyme mapping of vaccinia virus DNA.

The cleavage sites for the restriction enzymes Bg/I, HindIII, KpnI, SalI, SmaI, and XhoI were located, from primary data, on the DNA isolated from the WR strain of vaccinia virus. Bg/I and SmaI divide the DNA into five segments which can be isolated by sucrose gradient centrifugation. These large segments provide a convenient means to group segments produced by other enzymes. The construction of physical maps was initiated by identifying the segments at each end of the DNA and then finding segments which were adjacent to these terminal sections. This was accomplished by isolating large shear fragments which contained the covalently linked termini of the DNA. Most of the data needed to derive the maps were obtained by isolating segments produce by one enzyme and then cleaving these individual segments with a second enzyme.

Base Sequence↗
PubMed1996

A long-range physical map of human chromosome 21q22.1 band from the YAC continuum.

The human Chromosome (Chr) 21q22.1 region contains several genes for cytokines and neurotransmitters and the gene for superoxide dismutase (mutant forms of which can cause familial amyotrophic lateral sclerosis). A region of approximately 5.8 Mb encompassing D21S82 and the glycinamide ribonucleotide transformylase (GART) loci was covered by overlapping YAC clones, which were contiguously ordered by clone walking with sequence-tagged site (STSs). A total of 76 markers, including 29 YAC end-specific STSs, were unambiguously ordered in this 5.8-Mb region, and the average interval between markers was 76 kb. Restriction maps of the YAC clones with rare-cutting enzymes were simultaneously prepared, and the restriction sites were aligned to obtain a consensus restriction map of the proximal region of the 21q22.1 band. The restriction map made from 44 overlapping YACs contains 54 physically assigned STSs. By integrating the consensus map of the adjacent 1.8-Mb region, we obtained a fine physical map spanning 6.5 Mb of human Chr 21q22.1. This map contains 24 precisely positioned end-specific STSs and 12 NotI-linking markers. More than 39 potential CpG islands were identified in this region and were found to be unevenly distributed. This physical map and the YACs should be useful as a reference map and as a resource for further structural analysis of the Giemsa-negative band (R-band) of Chr 21q22.1.

Base Sequence↗
PubMed1984

Progressive multifocal leucoencephalopathy: analysis of JC virus DNA from brain and kidney tissue.

JC virus-specific DNA from both brain and kidney of the PML case G.S. was analysed by restriction mapping and Southern blot analysis. The genome of JCV DNA from brain (JCV GS-B) was full length, whereas JCV DNA GS from kidney (JCV GS-K) was 120 bp smaller. Restriction maps, constructed from JCV GS-B using various enzymes showed that most cleavage sites were consistent with standard map sites. Comparison of virus DNA from brain and kidney revealed that JCV GS-specific cleavage sites were identical, and that the DNAs were therefore of the same subtype. The deletion in kidney DNA was situated in the area of the origin of replication which is known to be hypervariable in the JCV genome. Restriction maps revealed that the DNA from each organ was homogeneous in length but a proportion of the DNA molecules were heterogeneous in restriction sites. It is concluded from these data that initial infection with one JCV subtype was followed by the development of heterogeneous DNA molecules.

Brain↗
PubMed1993

A cladistic analysis of phenotypic associations with haplotypes inferred from restriction endonuclease mapping. IV. Nested analyses with cladogram uncertainty and recombination.

We previously developed an analytical strategy based on cladistic theory to identify subsets of haplotypes that are associated with significant phenotypic deviations. Our initial approach was limited to segments of DNA in which little recombination occurs. In such cases, a cladogram can be constructed from the restriction site data to estimate the evolutionary steps that interrelate the observed haplotypes to one another. The cladogram is then used to define a nested statistical design for identifying mutational steps associated with significant phenotypic deviations. The central assumption behind this strategy is that a mutation responsible for a particular phenotypic effect is embedded within the evolutionary history that is represented by the cladogram. The power of this approach depends on the accuracy of the cladogram in portraying the evolutionary history of the DNA region. This accuracy can be diminished both by recombination and by uncertainty in the estimated cladogram topology. In a previous paper, we presented an algorithm for estimating the set of likely claodgrams and recombination events. In this paper we present an algorithm for defining a nested statistical design under cladogram uncertainty and recombination. Given the nested design, phenotypic associations can be examined using either a nested analysis of variance (for haploids or homozygous strains) or permutation testing (for outcrossed, diploid gene regions). In this paper we also extend this analytical strategy to include categorical phenotypes in addition to quantitative phenotypes. Some worked examples are presented using Drosophila data sets. These examples illustrate that having some recombination may actually enhance the biological inferences that may derived from a cladistic analysis. In particular, recombination can be used to assign a physical localization to a given subregion for mutations responsible for significant phenotypic effects.

Algorithms↗
PubMed1977

HindII, HindIII, and HpaI restriction fragment maps of the left arm of bacteriophage lambda DNA.

The sites on the left arm of bacteriophage lambda DNA cleaved by the restriction endonucleases isolated from Hemophilus influenzae strain Rc (HincII) and Rd (HindII + III), and Hemophilus parainfluenzae (HpaI) were localized on the lambda physical map, and the fragments resulting from these cleavages were identified by gel electrophoresis. The restriction sites within the b2 region of lambda were mapped by analysis of the digestion profiles of deletion and substitution derivatives of lambda, as well as by digesting individual fragments produced by one restriction endonuclease with another restriction endonuclease. The restriction sites of the lambda genome between the left vegetative end and the b2 region were mapped entirely by succesive digestion experiments. The restriction fragment map for the right arm of lambda may be found in the accompanying paper (Robinson and Landy, 1977).

Chromosome Mapping↗
PubMed1982

Antigen recognition by cloned cytotoxic T lymphocytes follows rules predicted by the altered-self hypothesis.

Radiation chimeras prepared by injecting H-2 heterozygous F1 stem cells into lethally irradiated parental hosts show a marked, but not absolute, preference for host-type H-2 antigens in the H-2-restricted cytotoxic T lymphocyte (CTL) response to minor histocompatibility (minor H) antigens. We have selected for the anti-minor HCTL that are restricted to the parental H-2 type absent from the chimeric host and found that in two out of eight cases, such CTL lysed target cells of either parental H-2 type. From one of these CTL populations that lysed H-2d and H-2k target cells expressing BALB minor H antigens, clones were derived and further analyzed. The results showed that: (a) lysis of both H-2d and H-2k target cells was H-2 restricted; (b) H-2d restriction mapped to Dd, and H-2k restriction mapped to Kk; (c) testing against various H-2d and H-2k strains of different and partially overlapping minor H backgrounds as well as against the appropriate F1 crosses revealed that in Dd- and Kk-restricted killing, different minor H antigens were recognized. In a second system, a CTL population was selected from normal (H-2d x H-2k)F1 mice that was specific for H-2d plus minor H antigens and for H-2k plus trinitrophenylated bovine serum albumin. We interpret these findings in terms of the altered-self hypothesis: The association of one H-2 antigen with one conventional antigen X may be recognized by the same T cell receptor specific for the complex formed by a different H-2 antigen in association with a second conventional antigen Y. The implications of these observations for the influence of self H-2 on the generation of the T cell receptor repertoire are discussed.

Animals↗
PubMed1991

Mapping sequenced E.coli genes by computer: software, strategies and examples.

Methods are presented for organizing and integrating DNA sequence data, restriction maps, and genetic maps for the same organism but from a variety of sources (databases, publications, personal communications). Proper software tools are essential for successful organization of such diverse data into an ordered, cohesive body of information, and a suite of novel software to support this endeavor is described. Though these tools automate much of the task, a variety of strategies is needed to cope with recalcitrant cases. We describe such strategies and illustrate their application with numerous examples. These strategies have allowed us to order, analyze, and display over one megabase of E. coli DNA sequence information. The integration task often exposes inconsistencies in the available data, perhaps caused by strain polymorphisms or human oversight, necessitating the application of sound biological judgment. The examples illustrate both the level of expertise required of the database curator and the knowledge gained as apparent inconsistencies are resolved. The software and mapping methods are applicable to the study of any genome for which a high resolution restriction map is available. They were developed to support a weakly coordinated sequencing effort involving many laboratories, but would also be useful for highly orchestrated sequencing projects.

Base Sequence↗
PubMed1983

Herpes simplex encephalitis: analysis of a cluster of cases by restriction endonuclease mapping of virus isolates.

In December 1979, there were three deaths from culture-proven herpes encephalitis in 3 weeks in the New Haven area, and a nurse caring for one of these patients developed a herpetic lesion on her nose. The three brain isolates, the isolate from the nurse, and several epidemiologically unrelated strains were analyzed by restriction endonuclease mapping. All were determined to be distinct strains of herpes simplex virus. The possibility that a single strain of virus caused this cluster of cases was therefore examined directly and disproved.

Adolescent↗
PubMed1990

Cloning and characterization of the rRNA genes and flanking regions from Babesia bovis: use of the genes as strain discriminating probes.

Three sets of rRNA genes (units 1S-3S) have been identified in Babesia bovis (Samford isolate). All three units are present in the same, probably single, copy number. The rRNA genes and flanking regions have been analysed by cloning, restriction mapping and DNA hybridization. The units are approximately 7 kb in length and have essentially identical restriction maps. In contrast the flanking regions exhibit significant restriction site differences. However, the regions upstream of all three units are related and sequences similar to part of the region upstream of units 1S and 3S are present in multiple copies in the genome. The downstream regions appear to be unrelated, but downstream from unit 1S is a region of at least 7 kb similar to a second region not closely linked to the rDNA units. The restriction enzyme site polymorphisms in the flanking regions of the equivalent units in different isolates allow ready discrimination among six different isolates of B. bovis.

Animals↗
PubMed1981

Map of restriction sites on bacteriophage T4 cytosine-containing DNA for endonucleases bamHI, BglII, KpnI, PvuI, SalI, and XbaI.

A complete map of the cleavage sites of restriction endonucleases BamHI, BglII, KpnI, PvuI, SalI, and XbaI was determined for the cytosine-containing DNA of a bacteriophage T4 alc mutant. The 56 sequence-specific sites were assigned map coordinates based on a least-squares analysis of measured fragment lengths. Altogether, the lengths of 118 fragments from single and double enzyme digestions were measured by electrophoresis of the fragments in agarose gels. DNA fragments of known sequence or DNA fragments calibrated with fragments of known sequence were used as standards. The greatest deviation between an experimentally measured fragment length and its computed map coordinates was 3.0%; the average deviation was 0.8%. The total length of the wild-type T4 genome was calculated to be 166,200 base pairs.

Chromosome Mapping↗
PubMed1993

Typing of human papillomavirus DNAs by restriction endonuclease mapping of the PCR products.

The polymerase chain reaction (PCR) for the diagnosis of human papillomavirus (HPV) infections, and in particular for the study of cervical HPV-associated lesions, is used widely. We identified a novel set of universal primers that are able to amplify a fragment spanning the E1 open reading frame (ORF) from different mucosotropic HPV types. A restriction endonuclease digestion of the amplified products is suggested for accurate typing. In particular, AluI digestion of the amplified fragments yields a distinctive fragment pattern for each 'high-risk' (16, 18, 31 and 33) HPV sequence, thus distinguishing them from 'low-risk' (6b and 11) HPV sequences.

Base Sequence↗
PubMed1990

Common antibiotic resistance plasmids in Staphylococcus aureus and Staphylococcus epidermidis from human and canine infections.

The plasmids of a multiresistant "canine" Staphylococcus epidermidis-culture were investigated. Two small plasmids, the 4.55 kB chloramphenicol resistance (CmR-) plasmid pSC4 and the 4.45 kB tetracycline resistance (TetR-) plasmid pST 3 could be isolated. Detailed restriction maps of pSC 4 and pST 3 were constructed by double restriction endonuclease digests. The restriction maps revealed extensive structural homologies between pSC 4 from "canine" S. epidermidis and the CmR-plasmid pC 221 from "human" S. aureus as well as between pST 3 from "canine" S. epidermidis and the TetR-plasmid pT 181 from "human" S. aureus. These data suggested that an exchange of small plasmids between S. epidermidis and S. aureus might be possible.

Animals↗
PubMed1977

Mapping of restriction sites in the attachment site region of bacteriophage lambda.

A find structure map of the EcoRI fragment containing the lambda attachment-site region has been constructed. 38 different restriction endonucleases have been employed and 170 sites located in this fragment. In addition, sites in adjacent regions have been determined for several enzymes. Complete cleavage maps of the entire lambda genome have been obtained for endonucleases BglII, BluI, KpnI, SacI, SacII, SalI and XbaI. The strategy employed for mapping included comparison of deletion and substitution mutants, analysis of mixed digests, and detailed analysis of subfragments.

Chromosome Mapping↗
PubMed1997

Detection and typing of human papillomavirus DNAs by restriction endonuclease mapping of the PCR products.

Human papillomavirus (HPV) has been implicated strongly with human cervical, anal and penile cancers. The polymerase chain reaction (PCR) was used to detect HPV in cervical specimens of 88 women working at the public whorehouse. Using consensus primers which encode the L1 region of the HPV genome, the presence of HPV DNA was demonstrated in 2 specimens. Restriction endonuclease digestion of the amplified products was carried out for accurate typing. Samples which were positive by L1 PCR were digested with Hae III, BstN I and Dde I restriction enzymes. The patterns produced by digestion were identified as HPV types 6b and 16.

Adult↗
PubMed1989

Restriction site mapping of adenovirus types 9 and 15 and genome types of intermediate adenovirus 15/H9.

Physical maps were established for seven genome types of the intermediate adenovirus (AV) 15/H9 and for the serologically related prototypes AV15 and 9 using the enzymes BamHI, BglII, and HindIII. The polarity of the fragment order was determined by hybridization with known AV2 fragments. AV15/H9 strains showed more restriction sites in common with AV9 than with AV15. The results are consistent with the hypothesis that intermediate AV15/H9 strains have emerged by recombination.

Adenoviruses, Human↗
PubMed1987

The genomic RNA of diabetogenic encephalomyocarditis virus: characterization and molecular cloning.

The RNA of a diabetogenic variant of encephalomyocarditis (EMC) virus (D variant, ifp- phenotype) and a nondiabetogenic variant of EMC virus (B variant, ifp+ phenotype) which were derived from the same virus stock (M strain) have been compared. The size of both genomes is estimated at 7.7 kb. The poly(C) tract of EMC-D is estimated at 144 bases, whereas that of EMC-B is 141 bases in length. The untranslated 5' terminal 103 nucleotides are identical for B and D with preservation of a stable terminal hairpin structure. The entire open reading frame of both variants has been cloned and the restriction maps of 12 different enzymes are identical. These maps were compared to a computer-generated restriction map of another strain of EMC virus which has been cloned and sequenced by Palmenberg et al. (1984, Nucleic Acids Res. 12, 2969-2985). Approximately 50% of the restriction sites of the B and D variants had similar locations in this strain of EMC. We conclude that significant genetic variation exists between the M strain (B and D variants) and the Palmenberg strain of EMC virus. However, the B and D variants are very similar at the molecular level and comparison of their nucleotide sequences will be necessary to reveal the basis for their different biological properties.

Animals↗
PubMed1986

The genome of bovine herpesvirus 1 (BHV-1) strains exhibiting a neuropathogenic potential compared to known BHV-1 strains by restriction site mapping and cross-hybridization.

Bovine herpesvirus 1 (BHV-1) strains can be differentiated by their DNA and polypeptide patterns, and by antigenic properties as demonstrated by monoclonal antibodies. We classified the BHV-1 strains according to these data as BHV-1.1, BHV-1.2 (a/b) and BHV-1.3 (a/b). BHV-1.1 and BHV-1.2 correspond to the well known 'common' BHV-1 strains, whereas BHV-1.3 has only recently been recognized and exhibits a neuropathogenic potential. In the present paper we describe the structural genome characteristics of BHV-1.3 compared to those of the other BHV-1 strains, examined by means of restriction site mapping, electron microscopy and cross-hybridization. Our results also confirm and complete data concerning BHV-1.1 and BHV-1.2 published by other authors. The following main conclusions can be drawn from our investigations: BHV-1.1 and BHV-1.2 differences are restricted to distinct genomic regions, characterized by loss or gain of restriction sites. BHV-1.3, however, differs from the other BHV-1 strains in restriction site alterations throughout the whole genome. Electron microscopy showed the typical BHV-1 DNA structure for BHV-1.3. Genetic homology between BHV-1.1 and BHV-1.2, reported to be about 95%, was confirmed by cross-hybridization, and a similar high base sequence homology for BHV-1.3 could be shown.

Animals↗