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M H Edgell

Publications and source records attributed to M H Edgell.

At least 73 records · Page 4Linked to original sources
PubMed1979

DNA fragments of the Mus musculus beta globin haplotypes Hbbs and Hbbd.

Two alternative haplotypes at the complex locus controlling hemoglobin beta chain synthesis in Mus musculus were compared at the DNA level. As expected, Hbbd homozygotes--which as adults synthesize two species of beta chain--have two genes for beta globin. Adult mice homozygous for the Hbbs haplotype make only a single type of beta polypeptide, yet they also have two beta globin genes. Apparently the two Hbbs genes encode identical proteins, or one of the two genes is not detectably expressed. The Hbbs and Hbbd haplotypes are thus more similar at the DNA level than studies of their polypeptide products have indicated.

Animals↗
PubMed1978

Mutagenesis at a specific position in a DNA sequence.

Predefined changes in a known DNA sequence were introduced by a general method. Oligodeoxyribonucleotides complementary to positions 582 to 593 of the viral DNA strand of the bacteriophage phiX174 am3 mutant (pGTATCCTACAAA), and to the wild type sequence in this region (pGTATCCTACAAA), were synthesized and used as specific mutagens. Each of these oligonucleotides was incorporated into a complete circular complementary strand when used as primer on a genetically heterologous viral strand template, by the combined action of subtilisin-treated Escherichia coli DNA polymerase I and T4 DNA ligase. Incomplete duplexes were removed or were inactivated by nuclease S1 and the products were used to transfect spheroplasts of E. coli. Both oligonucleotides induced specific mutations at high efficiency when used with heterologous template (15% mutants among progeny phage). The am phages isolated by this procedure are phenotypically gene E mutants, and contain A at position 587 of the viral strand. They thus appear identical with am3 and provide evidence that the change G leads to A at position 587 is sufficient to produce a defective E function. Since the template for the induction of am mutants carried another genetic marker (sB1), the strains carrying the induced mutations have the new genotype am3 sB1. It should be possible to introduce the am3 mutation into any known mutant strain of phi174 using this same oligonucleotide. Both possible transition mutations were induced in these experiments. In principle, the method could also induce transversions, insertions, and deletions. The method should be applicable to other circular DNAs of similar size, for example recombinant DNA plasmids.

Base Sequence↗
PubMed1978

A comparison of two cloned mouse beta-globin genes and their surrounding and intervening sequences.

The BALC/c mouse has two nonallelic beta-globin genes that appear to reside on two different Eco R1 fragments of genomic DNA. We have already cloned one of these fragments and shown that the gene encoded within it is interrupted by at least one large intervening sequence of DNA. We have now cloned and characterized the second beta-globin gene-containing fragment. The coding sequence of its gene is also interrupted by an intervening sequence of DNA that occurs in about the same position, relative to the coding sequence, as does the first. Because some shared features of the structure of these two genes might be responsible for their coordinate expression and the elimination of their intervening sequences, we have compared their surrounding, coding and intervening sequences by restriction endonuclease analysis and by visualization of the heteroduplex structures formed between them. Of the 7000 bp of sequence compared in this way, we find only a few hundred base pairs of homology in addition to the coding sequence. These shared sequences flank the coding sequence and appear to include only those portions of the intervening sequence immediately adjacent to the interrupted structural gene.

Animals↗
PubMed1978

Recognition site of Escherichia coli B restriction enzyme on phi XsB1 and simian virus 40 DNAs: an interrupted sequence.

Methyl groups placed on varphiXsB1 replicative form DNA by the Escherichia coli B modification enzyme are located in the overlap between fragments Mbo II-3 and Alu I-2, a 61-base-pair DNA segment. Mutations that led to loss of susceptibility to restriction by E. coli B occurred within this segment at three positions spanning 14 nucleotides. A sequence difference between varphiXsB1 and varphiXam3cs70, a varphiX174 strain not restricted by E. coli B, occurs at one of these positions. The site on simian virus 40 DNA methylated by the modification enzyme is located in the 115-base-pair overlap between fragments Hae III-I and Alu I-G. The sequences of these segments of varphiXsB1 and simian virus 40 DNA and two regions of phage f1 DNA recognized by the E. coli B restriction enzyme [Ravetch, J. V., Horiuchi, K. & Zinder, N. D. (1978) Proc. Natl. Acad. Sci. USA 75, 2266-2270] contain a homology of nine bases in the configuration:5'-T-G-A... 8N... T-G-C-T... 9N... T-N-N-T-3'. The sequence 5'-T-G-A... 8N... T-G-C-T-3' may constitute the restriction enzyme recognition site since it does not occur in varphiXam3cs70 DNA and occurs only once in simian virus 40 DNA, and since all observed mutations leading to loss of the site occur at one of the bases specified by this sequence. Analysis of the sequence of varphiXam3cs70 showed that if no other residues are recognized, all seven of these bases are essential for recognition and the interval between the two groups of specified bases must be precisely eight.

Base Sequence↗
PubMed1978

Multiple related immunoglobulin variable-region genes identified by cloning and sequence analysis.

We have identified at least six EcoRI fragments of mouse DNA that encode variable-region gene sequences closely related to the mouse kappa light chain, MOPC-149. Two of these fragments have been cloned, and the entire nucleotide sequence of the variable-region genes encoded on each has been determined. Both genes encode closely related variable-region sequences extending from codon position 1 through position 97. Neither fragment encodes a constant-region sequence. Although both genes are closely related, they differ from one another and from the sequence expressed in the MOPC-149 cell from which they were cloned. These few differences cluster within the complementarity-determining regions although several occur in framework sequences as well. We therefore conclude that an antibody-producing cell contains genetic information corresponding to its expressed sequence and several other closely related but silent sequences. These initial results raise the possibility that similar sets of genes might exist corresponding to each of the many subgroups already identified among mouse kappa light chains. If true, this would further suggest that the mouse genome might be rich enough in variable-region genes so as to encode a major portion of the variable-region repertoire.

Animals↗
PubMed1978

The ribosomal RNA gene region in the sea urchin Lytechinus variegatus.

A fine structure map of the ribosomal gene region of the sea urchin Lytechinus variegatus has been generated. The ribosomal DNA density satellite has been analyzed with restriction enzymes and by hybridization to urchin 18S and 26S ribosomal RNA (rRNA). The L. variegatus rRNA repeats show heterogeneity on the nontranscribed spacer. Minor bands containing transcribed spacer sequences are observed. One interpretation of these bands would require both transcribed spacer heterogeneity and defective ribosomal genes retained among the rRNA gene repeats.

Animals↗
PubMed1977

Isolation and characterization of the four major proteins in the virion of bacteriophage phiX174.

A preparative method is described for the isolation of the major protein species from the virion of bacteriophage phiX174. Two proteins, the cistron G and H products, are located in the virion spikes. After removal of the spikes, the capsid contains the cistron F product as well as a small protein which is the product of cistron J and the majority of the DNA. During the removal of the spikes, a precipitate containing the F and G proteins is formed. The proteins from the spike, capsid, or precipitate can be isolated on the basis of size by gel-filtration chromatography. The cistron G protein has an aminoterminal methionine, while the small J protein has an amino-terminal serine. Amino acid compositions as well as peptide maps indicate each species is unique and that, in sum, they account for over half the coding capacity of the viral genome.

Amino Acids↗
PubMed1977

Amino acid sequence of the small core protein from bacteriophage phiX174.

The amino acid sequence of small core protein of bacteriophage phiX174 has been determined by a combination of automated Edman degradation of the intact polypeptide and by analysis of tryptic and thermolytic peptides. The six lysyl and six arginyl residues of this 37-residue polypeptide are concentrated in two structurally homologous 12-residue segments of the sequence. The hydrophobic residues of valine, tryptophan, tyrosine, and phenylalanine are contained in the carboxyl-terminal nine residues of the protein, together with one of the two leucyl residues and two of the three glutaminyl residues. The single free carboxyl group in the protein is the alpha-COOH of the C-terminal phenylalanyl residue. The overall sequence of this small core protein suggests that it may function as a DNA-condensing protein. The protein sequence presented here corresponds exactly to the DNA base sequence of the cistron J region of the phiX174 genome determined in another laboratory.

Amino Acid Sequence↗
PubMed1977

Cloning specific segments of the mammalian genome: bacteriophage lambda containing mouse globin and surrounding gene sequences.

We have developed a general approach to the cloning of specific segments of the mammalian genome that involves a two-step purification of EcoRI fragments of mammalian DNA and their in vitro insertion into a suitably constructed EK2 derivative of bacteriophage lambda. The combination of fragment purification, exclusion of parental-type recombinants, and simple phage screening techniques permits the isolation of virtually any gene segment for which there is an identifying hybridization probe. We illustrate the approach by describing the cloning of an approximately 7000-base-long segment of mouse DNA containing globin and surrounding gene sequences.

Animals↗
PubMed1975

Specific cleavage analysis of mammalian mitochondrial DNA.

Mitochondrial DNA from several mammalian species has been digested with a site-specific restriction endonuclease (HaeIII) from Haemophilus aegyptius. A quantitative analysis of the resulting specific fragments indicates that the mtDNA of any individual mammal is predominantly a single molecular clone. Gel analysis of specific cleavage products has proven quite sensitive in detecting differences in mtDNA: mtDNAs from the more distantly related mammals studied (e.g., donkey and dog) are found to have few bands in common and very closely related mammals (e.g., donkey and horse) share only about 50% of their bands. This procedure has detected several intraspecies mtDNA differences. Six distinct human patterns have been found, with one pattern usually differing from another in two or three bands. mtDNAs from different organs of single individuals have also been analyzed, and no differences have been found.

Animals↗
PubMed1973

Mapping of new Escherichia coli K and 15 restriction sites on specific fragments of bacteriophage phi X174.

We have isolated several new phiX174 mutants which contain sites sensitive to restriction by Escherichia coli. One contains an E. coli 15 restriction site and three are double mutants containing an E. coli K site as well as the E. coli 15 site. The replicative form (RF) DNA of one of the mutants containing a K site has been shown to be restricted in spheroplasts of a K-12 strain. The infectivity of this RF, but not wild-type RF, has also been shown to be inactivated by an E. coli K extract and by purified K restriction enzyme in vitro. The product of the RF treated with purified K restriction enzyme in vitro is a full length linear molecule. The mutant sites have also been localized to specific regions of the phiX174 genome by a fragment mapping technique, making use of specific fragments of phiX174 RF DNA obtained by digestion with a specific endonuclease.

Chromosome Mapping↗
PubMed1972

Specific fragments of phi X174 deoxyribonucleic acid produced by a restriction enzyme from Haemophilus aegyptius, endonuclease Z.

A restriction-like enzyme has been purified from Haemophilus aegyptius. This nuclease, endonuclease Z, produces a rapid decrease in the viscosity of native calf thymus and H. influenzae deoxyribonucleic acids (DNA), but does not degrade homologous DNA. The specificity of endonuclease Z is different from that of the similar endonuclease isolated from H. influenzae (endonuclease R). The purified enzyme cleaves the double-stranded replicative form DNA of bacteriophage phiX174 (phiX174 RF DNA) into at least 11 specific limit fragments whose molecular sizes have been estimated by gel electrophoresis. The position of these fragments with respect to the genetic map of phiX174 can be determined by using the genetic assay for small fragments of phiX174 DNA.

Ammonium Sulfate↗