Showing papers on "genomic DNA published in 1978"

Cloning human fetal γ globin and mouse α-type globin DNA: Preparation and screening of shotgun collections

Abstract: Shotgun collections of Charon 3A bacteriophages containing Eco RI fragments of human and mouse DNA were constructed with the use of in vitro packaging. Plaques were screened by hybridization, and globin-specific clones were isolated from both human (Charon 3AHs51.1) and mouse (Charon 3AMm30.5). The fragments cloned were detected in unfractionated genomic DNA by the Southern method of hybridization.

Intragenic DNA spacers interrupt the ovalbumin gene

Abstract: We have performed restriction endonuclease mapping to examine the linear organization of the ovalbumin gene in chromosomal DNA. Treatment of genomic DNA with restriction endonucleases that do not cleave the ovalbumin mRNA sequence results in the generation of multiple DNA fragments capable of annealing with ovalbumin-specific probes in molecular hybridization reactions. These data strongly suggest that the linear order of DNA sequences coding for ovalbumin is interrupted by at least two intragenic DNA spacers absent from the corresponding RNA. At least one of these spacer sequences interrupts the coding sequence; therefore the chromosomal ovalbumin gene is not colinear with its translational product. We can discern no difference in the sequence organization about this gene in producer and nonproducer somatic cells, suggesting that the presence of intragenic spacers does not reflect that transcriptional activity of this gene. Furthermore, profiles obtained for gametes are identical to those observed for somatic cells, indicating that significant translocation during development is not responsible for the generation of this split sequence. Divergence of the intragenic spacer, however, is observed between individual chickens; thus multiple alleles may exist for this gene that are identifiable by differences in the organization of spacer with no apparent phenotypic evidence for their presence.

The Sequence of the Chromosomal Mouse /3-Globin Major Gene: Homologies in Capping, Splicing and

Abstract: David A. Konkel, Shirley M. Tilghman* and Philip Leder Laboratory of Molecular Genetics National Institute of Child Health and Human Development Bethesda, Maryland 20014 Summary We have determined the entire nucleotide se- quence of a cloned j3-globinma’ gene derived from the BALB/c mouse. This sequence is 1567 bases long and includes the 5’ cap region as well as the presumptive poly(A) addition site of @globin mRNA. The sequence establishes the fact that the gene is encoded in three discontinuous seg- ments of DNA interrupted by two intervening se- quences and precisely locates each. The smaller intervening sequence, 116 bases long, occurs between Arg and Leu codons at codon positions 30 and 31. The larger intervening sequence of 646 bases also occurs between Arg and Leu codons, but at codon positions 104 and 105. There is striking homology between the borders of the two intervening sequences, but no extensive dyad symmetry. Furthermore, the DNA region that just precedes and overlaps the 5’ cap structure of the mRNA shows homology to corresponding regions in other eucaryotic genes including the late ade- novirus promoter. The 3’ untranslated sequence is closely homologous to that of the rabbit p- globin mRNA. The sequence thus allows us to identify several noncoding regions of potential importance for the expression and processing of genetic information. It also provides a basis for future comparison with other sequenced genes and a defined substrate for the development of direct tests of gene function. The adult BALB/c mouse coordinately expresses two P-globin genes encoding the two polypeptides /3-globinmaJ and @globin”‘“. These genes reside on the two Eco RI fragments of genomic DNA that have been cloned in the bacteriophage A (Tilghman et al., 1977; Tiemeier et al., 1978). Both genes appear to be interrupted by two intervening se- quences of DNA (ibid.) that are transcribed into the 1% globin mRNA precursor (Smith and Lingrel, 1978; Kinniburgh, Mertz and Ross, 1978; Tilghman et al., 1978b). Such structures have also been

DNA sequence organization in the common Pacific starfish Pisaster ochraceous

Abstract: The sequence arrangement of the genomic DNA from the common sea star Pisaster ochraceous has been examined. Reassociation kinetics at DNA fragment lengths of 300 base pairs (bp) indicate the presence of at least three repetitive components in this DNA. The majority of these repetitive sequences are reiterated over the range from 10 to 100's. Approximately one-third of the nucleotides are found in repetitive sequences. Analysis of the reassociation kinetics of 3000-bp DNA fragments demonstrates the interspersion of repetitive and unique DNA sequences. The hyperchromicity of 3000-bp fragments reassociated to low Cot values (the product of moles of nucleotide per litre and time in seconds), and the size distribution of S1 nuclease resistant DNA duplex in these reassociation products, indicate a short-period interspersion pattern in the starfish genome. Repetitive segments (400 +/- 100 bp) are interspersed with longer unique DNA sequences. At a fragment length of 3000 bp the major fraction of the single-copy DNA is found in such an arrangement. In addition to short repetitive segments a substantial portion of the repetitive DNA nucleotides are found in segments excluded by Sepharose CL-2B (greater than or equal to 2000 bp). As much as one-quarter of the repetitive sequence nucleotides can be assigned to long segments.

Amplification and characterization of eukaryotic structural genes.

Abstract: An approach to the study of eukaryotic structural genes which are differentially expressed during development is described. This approach involves the isolation and amplification of mRNA sequences by in vitro conversion of mRNA to double-stranded cDNA followed by molecular cloning in bacterial plasmids. This procedure provides highly specific hybridization probes that can be used to identify genes and their contiguous DNA sequences in genomic DNA, and to detect specific RNA transcripts during development. The nature of the method allows the isolation of individual mRNA sequences from a complex population of molecules at different stages of development.

Gene Expression in Normal and Neoplastic Breast Tissue

Abstract: Most structural gene sequences in mammalian cells represent less than one millionth of the information contained in the genomic DNA. Because of this enormous complexity of genetic information in higher organisms, an understanding of the mechanisms regulating gene expression requires the study of specific genes. Fortunately, considerable progress has been made in the last decade in the development of techniques for the isolation of individual eukaryotic messenger RNAs (mRNAs) and the synthesis of their complementary DNA copies (cDNAs) (for a general review, see Rosen and Monahan(1)). These molecular hybridization probes have been utilized successfully to study gene expression in a number of model systems, notably the control of globin gene expression during erythroid differentiation(2) and steroid hormone induction of ovalbumin mRNA(3) and the other egg-white protein mRNAs(4) in the chick oviduct. These studies have recently been culminated by the determination of the entire nucleic acid sequence of the rabbit s-globin mRNA.(5) However, the precise mechanism controlling the expression of any single eukaryotic gene remains to be established.