Showing papers on "genomic DNA published in 1983"
TL;DR: It is suggested that, in the eight lines which have amplified NB-19-21, the amplification units are overlapping, but not identical, and that transposition of the common sequences may occur prior to amplification.
653 citations
TL;DR: The molecular cloning and characterization of the gene encoding exo–cellobiohydrolase I of Trichoderma reesei strain L27 is reported and it is found to contain a 10 bp sequence, CAGCT–GACTG, that is homologous to a sequence necessary for splicing of introns in yeast2.
Abstract: The molecular cloning and characterization of the gene encoding exo–cellobiohydrolase I (CBHI) of Trichoderma reesei strain L27 is reported. Two adjacent HindIII genomic fragments of 1.16 kb and 2.3 kb were identified using differential hybridization techniques and were sub–cloned into plasmid pBR322. The identification of the gene encoding CBHI was determined by hybrid selection and confirmed by DNA sequence analysis. There are two introns in the genomic DNA that were identified by comparing the coding sequence with the published amino acid sequence1 and confirmed by sequencing of cDNA clones. Both introns were found to contain a 10 bp sequence, CAGCT–GACTG, that is homologous to a sequence necessary for splicing of introns in yeast2.
408 citations
TL;DR: Nucleotide sequence analysis of the cloned DNAs has revealed the primary structures of the calf and human AChR α-subunit precursors, which exhibit marked sequence homology with their Torpedo counterpart.
Abstract: The nicotinic acetylcholine receptor (AChR) from fish electric organ is well characterized and is known to consist of five subunits present in a molar stoichiometry of α2βγδ (reviewed in refs 1–3). The mammalian skeletal muscle AChR is thought to have a similar subunit structure4–6. We have recently elucidated the primary structures of the α-, β-, γ- and δ-subunit precursors of the Torpedo californica AChR by cloning and sequencing cDNAs for these polypeptides7–9; cDNA sequences for the γ-subunit precursor of the T. californica AChR10 and the α-subunit precursor of the Torpedo marmorata AChR11,12 have also been reported by other groups. The four subunits exhibit conspicuous sequence homology and are similar in hydrophilicity profile and predicted secondary structure, thus being most probably oriented in a pseudosymmetric fashion across the membrane. The transmembrane topology of the subunit molecules and the locations of functionally important regions, such as the acetylcholine binding site and the transmembrane segments which may be involved in the ionic channel, have been proposed. We have now cloned cDNA for the α-subunit precursor of the calf skeletal muscle AChR and a human genomic DNA segment containing the corresponding gene. Nucleotide sequence analysis of the cloned DNAs has revealed the primary structures of the calf and human AChR α-subunit precursors, which exhibit marked sequence homology with their Torpedo counterpart. The protein-coding sequence of the human AChR α-subunit precursor gene is divided by eight introns into nine exons, which seem to correspond to different structural and functional domains of the subunit precursor molecule.
405 citations
TL;DR: Genomic DNA blots probed with isotype-specific subclones under stringent conditions demonstrate that the alpha-skeletal and alpha-cardiac muscle actin genes are single copy, whereas the cytoskeletal actins, beta and gamma, are present in multiple copies in the human genome.
Abstract: We have constructed isotype-specific subclones from the 3' untranslated regions of alpha-skeletal, alpha-cardiac, beta-cytoskeletal, and gamma-cytoskeletal actin cDNAs. These clones have been used as hybridization probes to assay the number and organization of these actin isotypes in the human genome. Hybridization of these probes to human genomic actin clones (Engel et al., Proc. Natl. Acad. Sci. U.S.A. 78:4674-4678, 1981; Engel et al., Mol. Cell. Biol. 2:674-684, 1982) has allowed the unambiguous assignment of the genomic clones to isotypically defined actin subfamilies. In addition, only one isotype-specific probe hybridizes to each actin-containing gene, with a single exception. This result suggests that the multiple actin genes in the human genome are not closely linked. Genomic DNA blots probed with these subclones under stringent conditions demonstrate that the alpha-skeletal and alpha-cardiac muscle actin genes are single copy, whereas the cytoskeletal actins, beta and gamma, are present in multiple copies in the human genome. Most of the actin genes of other mammals are cytoplasmic as well. These observations have important implications for the evolution of multigene families.
361 citations
TL;DR: The DNA sequences of a complementary deoxyribonucleic acid (cDNA) and a portion of the gene coding for human prothrombin have been determined and it is proposed that the leader sequence consists of a signal sequence and a pro sequence for the mature protein that circulates in plasma.
Abstract: The DNA sequences of a complementary deoxyribonucleic acid (cDNA) and a portion of the gene coding for human prothrombin have been determined. The cDNA was 2005 base pairs in length and was found to code for part of a leader sequence of 36 amino acids, 579 amino acids present in the mature protein, a stop codon, a noncoding region of 97 base pairs, and a poly(A) tail of 27 base pairs. It is proposed that the leader sequence consists of a signal sequence and a pro sequence for the mature protein that circulates in plasma. The 10 glutamic acid residues that are present in the amino-terminal region of prothrombin and are converted to gamma-carboxyglutamic acid in the mature protein are coded by only the GAG codon. The cDNA for prothrombin was also employed as a probe for screening a human fetal liver genomic DNA library. One of the strongly positive phage containing a human DNA insert of 5 kilobases was mapped with restriction endonucleases and sequenced. This DNA contained approximately half of the gene for human prothrombin and included six introns and five exons coding for amino acid residues 144-448. The two largest intervening sequences in the genomic DNA contained two copies each of AluI repetitive DNA.
294 citations
TL;DR: Analysis of three overlapping cDNA clones revealed most of the nucleotide sequence of AFP mRNA, and the remaining nucleotides at the 5' end of the mRNA were elucidated from a cloned genomic DNA fragment.
Abstract: DNA complementary to human alpha-fetoprotein (AFP)mRNA was cloned in the plasmid pBR322. Analysis of three overlapping cDNA clones revealed most of the nucleotide sequence of AFP mRNA, and the remaining nucleotides at the 5' end of the mRNA were elucidated from a cloned genomic DNA fragment. The amino acid sequence was deduced from the nucleotide sequence, which revealed 19 amino acids in the signal sequence and 590 amino acids in mature AFP. There are 15 regularly spaced disulfide bridges, which generate a folding structure having three repeating domains. There is one potential N-glycosylation site, Asn-Phe-Thr, in the amino acid sequence. In comparison with mouse AFP, 66% of the amino acid sequence was conserved, with the highest identity (72%) in domain 3, followed by domain 2 (67%) and domain 1 (59%). In comparison with human albumin, a 39% conservation of primary structure was found. Again, the similarity was the highest in domain 3 and the lowest in domain 1. Human AFP and human albumin are similar in overall structure, but certain parts of the molecules differ significantly in their predicted secondary structure.
292 citations
TL;DR: Using a combination of chromosome walking and jumping, a DNA region from Drosophila containing Antennapedia is cloned, and the unusual size and complexity of this locus are discussed.
Abstract: Homeotic genes are involved in the control of developmental pathways: dominant mutations at the Antennapedia locus of Drosophila, for example, lead to replacement of the antennae on the head of the fly by mesothoracic legs. Using a combination of chromosome walking and jumping, we have cloned a DNA region from Drosophila containing Antennapedia. Five DNA inversion rearrangements which are associated with the Antennapedia mutant phenotype were localized within a 25-kb region. Genomic DNA sequences from this area were used as hybridization probes to screen cDNA libraries prepared from Drosophila embryonic and pupal poly(A)+ RNA. A 2.2-kb cDNA sequence (903) was isolated which appears to derive from at least four non-contiguous chromosomal regions that span 100 kb. It includes the positions of the inversion breakpoints. A second cDNA of 2.9 kb (909) is composed of sequences from at least three chromosomal regions, two of which are similar or identical to sequences contained in the 903 clone but the third is derived from genomic DNA within a putative 903 intron. The unusual size and complexity of this locus are discussed.
239 citations
TL;DR: This chapter discusses the use of Xenopus oocytes for the expression of cloned genes, one of the first systems in which purified DNA was correctly transcribed and expressed as protein.
Abstract: Publisher Summary This chapter discusses the use of Xenopus oocytes for the expression of cloned genes. The injection of amphibian oocytes was one of the first systems in which purified DNA was correctly transcribed and expressed as protein. An amphibian oocyte is a single large cell, surrounded by several thousand small follicle cells. It is in meiotic prophase, and active in RNA and protein synthesis, but totally inactive in DNA synthesis. The transcription of injected DNA takes place only in the nucleus or germinal vesicle of an oocyte, which is not normally visible. Gene isolation usually requires at least partially pure mRNA for the preparation of cDNA or for screening a genomic DNA library directly. As oocytes were first used for translating mRNA, cell-free systems have been greatly improved and generally have a lower background than oocytes.
233 citations
TL;DR: Screening a bank of mouse genomic DNA, cloned in Charon 4A, indicates that the number of actin-related sequences in the mouse genome is much higher than 20, and five phages have been isolated representing part of a sub-family of 20 to 50 similar but non-identical sequences, only weakly homologous to actin cDNA probes.
205 citations
TL;DR: The entire human pepsinogen C gene has been isolated from a cosmid genomic library and the nucleotide sequences of all the exons and the 5'- and 3'-flanking regions of the gene are presented, allowing us to conclude that the genes of these aspartic proteinases including peps inogen C are derived from a common ancestral gene.
181 citations
TL;DR: The gene that codes for the surface antigen of Plasmodium knowlesi sporozoites (CS protein) is unsplit and present in the genome in only one copy, indicating that the parasite promoter and ribosome binding site signals can be recognized in E. coli.
TL;DR: A DNA polymorphism between the Bristol and Bergerac strains has been used as a phenotypic marker in genetic crosses to map the cluster of actin genes within a 2% recombination interval on linkage group V between unc-23 and sma-1 in order to begin a molecular genetic analysis of the actin loci.
TL;DR: The findings show that IAP genetic elements can appear in new locations in mouse cellular DNA and suggest that this may occur through a process of proviral insertion.
Abstract: We analyzed two functionally defective mouse kappa light chain gene variants previously shown to contain novel insertions of repetitive DNA in their intervening sequences [Hawley, R. G., Shulman, M. J., Murialdo, H., Gibson, D. M. & Hozumi, N. (1982) Proc. Natl. Acad. Sci. USA 79, 7425-7429]. Heteroduplex analysis of the cloned genes shows that the insertions consist of intracisternal A-particle (IAP) genetic elements. Each insertion includes an IAP 5' long terminal repeat (LTR) sequence and extends to a characteristic IAP internal BamHI site where the IAP sequence is interrupted because the mutant genes were cloned from complete BamHI digests of the cellular DNAs. Restriction enzyme mapping indicates that the 5' LTR boundaries of the inserted IAP elements correspond closely to the previously determined rearrangement sites in the mutant genes. The IAP insertions in the two mutants can be distinguished by restriction-site differences and by the fact that one of them contains a deletion that is absent in the other. Nucleotide sequence data are presented for the LTRs of one full-length IAP gene copy randomly selected from a mouse genomic DNA library. These LTRs show many features typical of known integrated retroviral terminal repeat units, and the entire gene is bracketed by short direct repeats within the adjacent cellular DNA. Thus, the findings show that IAP genetic elements can appear in new locations in mouse cellular DNA and suggest that this may occur through a process of proviral insertion.
TL;DR: A synteny between myosin heavy chain and two unrelated markers, thymidine kinase and galactokinase, was found to be preserved in the rodent and human genomes.
Abstract: Cloned myosin heavy chain DNA probes from rat and human were hybridized to restriction endonuclease digests of genomic DNA from somatic cell hybrids and their parental cells. The mouse myosin heavy chain genes detectable by this assay were located on chromosome 11, and three different human sarcomeric myosin heavy chain genes were mapped to the short arm of chromosome 17. A synteny between myosin heavy chain and two unrelated markers, thymidine kinase and galactokinase, was found to be preserved in the rodent and human genomes.
TL;DR: The nucleotide sequences of the cDNA and genomic DNA clones permitted us to deduce the entire primary structure of the E beta chain and the complete exon-intron structure ofThe E beta gene.
Abstract: Using the cross-hybridization with plasmid pDC beta-1, containing the cDNA coding for the DC beta chain of the human major histocompatibility complex class II molecules, we have cloned and subjected to sequence analysis both the cDNA and genomic gene for the E beta chain of the BALB/c (d haplotype) mouse. The nucleotide sequences of the cDNA and genomic DNA clones permitted us to deduce the entire primary structure of the E beta chain and the complete exon-intron structure of the E beta gene. Unlike alpha chain genes that contain five exons, the E beta gene consists of six exons corresponding to the six functional domains--the leader, beta 1 and beta 2 domains, transmembrane peptide, intracytoplasmic peptide, and 3' untranslated region. In addition, two short blocks of sequences common to alpha and beta chain genes were identified in the 5' flanking regions. We propose that these sequences are involved in the coordinate expression of alpha and beta chains.
TL;DR: An understanding of the genetic relationship of the c-ras genes and additional genetic loci possibly involved in neoplastic transformation would be greatly facilitated by placement of the ras genes on the human chromosome map.
Abstract: Cellular transforming genes (c-onc) are evolutionarily conserved vertebrate DNA segments which have been identified by two different approaches. One group of these cellular genes has been defined by their close homology to the transforming genes of the acute transforming retroviruses (v-onc)1–3. The second group, which represent activated forms of normal cellular genes1,4–9, has been detected by the ability of certain genes from animal and human tumours to induce focal transformation of tissue culture cells. Investigation of the possibility that the same cellular gene might have given rise to both a retroviral and a tumour transforming gene revealed that two of the c-onc genes identified by transfecting genomic DNA from human tumours to murine 3T3 fibroblasts were related to the transforming genes of two closely related acute transforming retroviruses, Harvey murine sarcoma virus (HaMuSV) and Kirsten murine sarcoma virus (KiMuSV)10–12. The transforming genes of HaMuSV and KiMuSV are derived from two members of a cellular onc gene family called ras, which is a rather divergent group of normal vertebrate genes originally found by analysis of the cellular homologues of the v-onc genes of HaMuSV and KiMuSV13. Four distinct human cellular homologues of v-Ha-ras and v-Ki-ras (designated c-Ha-ras and c-Ki-ras, respectively) have been characterized14; two (c-Ha-ras-1 and c-Ha-ras-2) are more closely related to v-Ha-ras, while the others (c-Ki-ras-1 and c-Ki-ras-2) are more closely related to v-Ki-ras. On ligation with a retroviral long terminal repeat, the c-Ha-ras-1 gene of both rat and human have been shown to induce in vitro transformation of mouse NIH 3T3 cells by DNA transfection15,16. This gene and c-Ki-ras-2 have also been isolated as activated transforming genes in human tumours10–12. An understanding of the genetic relationship of the c-ras genes and additional genetic loci possibly involved in neoplastic transformation would be greatly facilitated by placement of the ras genes on the human chromosome map. Using DNA analysis of rodent×human somatic cell hybrids, we have now assigned each of the human genes to a different chromosome.
TL;DR: Results indicate that zein genes contain methylated sites that are demethylated in correlation with their expression and may be subject to rearrangements and transpositions.
TL;DR: A recombinant phage library is constructed that is enriched for DNA present in the HSR of this chromosome by using fluorescence-activated flow sorting for initial chromosome purification and demonstrated that cloned HSR segments were localized in the short arm of chromosome 2 in both normal and IMR-32 cells.
Abstract: Human neuroblastoma IMR-32 cells have large homogeneously staining regions (HSRs), primarily in the short arms of chromosome 1. We have constructed a recombinant phage library that is enriched for DNA present in the HSR of this chromosome by using fluorescence-activated flow sorting for initial chromosome purification. Eleven distinct cloned DNA segments were identified that showed significantly greater hybridization to IMR-32 genomic DNA, detected by Southern blotting, than to normal human genomic DNA. These sequences have also been localized to the HSR of chromosome 1 by in situ hybridization. Based on an approximate 50-fold sequence amplification for each cloned segment and a total HSR size of 150,000 kilobases, the amplified unit in the HSR is estimated to be 3,000 kilobases. Sequences homologous to all cloned HSR DNA segments were mapped to human chromosome 2 by using human-mouse hybrid cells. Further work using in situ hybridization demonstrated that cloned HSR segments were localized in the short arm of chromosome 2 in both normal and IMR-32 cells. Thus, the amplification of these sequences in IMR-32 cells may have involved transposition from chromosome 2 to chromosome I.
TL;DR: Results suggest that protein C23 has a preference for binding DNA sequences in the nontranscribed spacer of rDNA.
Abstract: Protein C23, a predominant nucleolar phosphoprotein and a putative nucleolus organizer protein, was analyzed for its general DNA binding characteristics and for its selectivity in binding plasmid DNAs containing cloned fragments of the genes that code for ribosomal RNA (rDNA). By use of nitrocellulose filter disk assays, the protein bound saturably to nuclear DNA with a relatively high affinity. Binding was maximal at low ionic strength (0-0.1 M KCl) with progressively decreasing binding at or above 0.2 M. In competition assays protein C23 showed a marked preference for linear single-stranded vs. double-stranded DNA and little or no affinity for ribosomal RNA. The relative affinities of rDNA sequences for protein C23 were determined with cloned fragments spanning 15.8 kilobases (kb) of DNA starting approximately 3.7 kb upstream from the initiation site for 45S preribosomal RNA to near the 3' end of the sequence coding for 28S RNA. Of the five linearized plasmids tested, only one (pKW1) was an effective competitor for 32P-labeled nuclear DNA. As measured by the concentration of competing DNA required to achieve 50% competition, pKW1 was approximately 20-fold more effective than the second best competitor. The DNA insert in pKW1 is a 3.5-kb sequence which is located in the nontranscribed spacer region less than 0.5 kb upstream from the initiation site for 45S preribosomal RNA. These results suggest that protein C23 has a preference for binding DNA sequences in the nontranscribed spacer of rDNA.
TL;DR: Analysis ofPublished data on cell size, maximum division rate, and genomic DNA content were compiled for over 70 species of unicellular prokaryotes and eukaryotes revealed statistically significant, positive correlations between cell volume and DNA content.
Abstract: Published data on cell size, maximum division rate, and genomic DNA content were compiled for over 70 species of unicellular prokaryotes and eukaryotes. Analysis of these data revealed statistically significant, positive correlations between cell volume and DNA content for both prokaryotes and eukaryotes, and between minimum cell doubling time and DNA content for eukaryotes. The relations exhibited by prokaryotes and eukaryotes are different from each other. The doubling time-DNA relation for unicellular eukaryotes is indistinguishable from a similar relation exhibited by plant root cells. Ciliates, which exhibit nuclear dualism, display relations between cell volume and total DNA content and between maximum division rate and micronuclear DNA content which are indistinguishable from those exhibited by uninucleate, unicellular eukaryotes.
TL;DR: The reduced methylation of the tumor and DNA correlated with the recent observation of other laboratories that individual genes are undermethylated in human cancer cells and that a number of different carcinogens can lower DNA methylation directly.
Abstract: The total genomic DNA methylation, i.e., the percentage of methylated cytosines, was measured in 20 cell lines derived from different types of human tumors. The measurements were obtained by cation-exchange liquid chromatography of bases released by formic acid hydrolysis. These experiments were done to determine if altered methylation is a prevalent and large defect in oncogenic transformation. A majority of the tumor cells measured had decreased levels of methylated DNA in comparison to our laboratory's and other laboratories' published measurements of normal cells and tissues. In fact, tumor cell DNA ranged as low as 1.2% of cytosines methylated compared to a value of 3% or more for normal cells and tissues. HpaII and MspI DNA restriction enzyme analysis confirmed for all tumor cell lines tested that their DNA was hypomethylated in comparison to the DNA from normal diploid fibroblasts tested. The results obtained by liquid chromatography and restriction enzyme analysis were strikingly similar. The reduced methylation of the tumor and DNA correlated with the recent observation of other laboratories that individual genes are undermethylated in human cancer cells and that a number of different carcinogens can lower DNA methylation directly.
TL;DR: Two mouse genomic DNA sequences homologous with human corticotropin‐β‐lipotropin precursor gene sequences have been cloned and a pseudogene corresponds to the functional mouse gene sequence encoding the carboxy‐terminal 143 amino acid residues and the 3′‐untranslated region.
TL;DR: The structure of the genomic mitochondrial sequence homologues indicates that during sea urchin evolution there occurred a germ-line transposition of a fragment of the mitochondrial genome into the nuclear DNA, followed by rearrangements and single nucleotide substitutions.
TL;DR: Comparison with pro-opiomelanocortin mRNA sequences from other species shows regions of high homology not only in the coding sequences but also in the 5'untranslated region where the first 50 nucleotides are over 80% purines.
Abstract: Polyadenylated RNA isolated from porcine pituitary neurointermediate lobes was used to construct a cDNA library The library was screened with a rat genomic DNA fragment specific for pro-opiomelanocortin sequences Two positive clones, pJA-19 and pJA-20, containing respectively 850 bp and 550 bp were characterized Sequence analysis of the cDNA inserts revealed the complete structure of the porcine pro-opiomelanocortin mRNA This mRNA would include 129 5'-untranslated nucleotides, 801 nucleotides coding for the 267 amino acids precursor and 162 3'-untranslated nucleotides Comparison with pro-opiomelanocortin mRNA sequences from other species shows regions of high homology not only in the coding sequences but also in the 5'untranslated region where the first 50 nucleotides are over 80% purines
TL;DR: The results indicate that the infected cells are of clonal origin with respect to the virus integration sites and they express multiple viral mRNA species including a 35S RNA.
Abstract: Human T-cell leukemia-lymphoma virus (HTLV) is the first unequivocal human retrovirus. Seroepidemiological and virus isolation studies indicate that HTLV is etiologically associated with a subtype of adult T-cell malignancy. We have molecularly cloned approximately 1 kilobase of sequences derived from the 5' and 3' termini of the HTLV genome. Use of these clones as probes allowed isolation of a 9.8-kilobase EcoRI fragment from a genomic DNA library of an HTLV-infected neoplastic T-cell line (CR). Analysis of this clone revealed the presence of cellular sequences flanking approximately 5 kilobases of viral sequences including one long terminal repeat sequence. The 5' and 3' clones, as well as subclones derived from different regions of the genomic clone, were used as probes to compare integrated proviruses and viral RNA expression in different HLTV-infected neoplastic T cell lines. The results indicate that the infected cells are of clonal origin with respect to the virus integration sites and they express multiple viral mRNA species including a 35S RNA.
TL;DR: The results of restriction endonuclease and hybridization analysis of DNA isolated from recombinant phages, and subcloned fragments thereof, were compatible with the restriction maps derived from digestion and Southern blot hybridization of genomic DNA.
Abstract: African trypanosomes are the causative agents of many medically and economically important diseases of man and domestic animals. The cell body of these blood-dwelling protozoa is enveloped with a dense layer of pellicular microtubules, which confer both motility and mechanical stability on these cells; microtubules are also important components of the flagellum. The major structural components of the microtubuli are two related proteins, alpha- and beta-tubulin. We have analyzed the genomic organization of alpha- and beta-tubulin genes in Trypanosoma brucei. In this organism, the majority of these genes are arranged in a single, tightly packed cluster of alternating alpha- and beta-tubulin genes with a basic repeat length together of 3.6 kilobase pairs. A genomic library of T. brucei was constructed by using the phage vector A 1059, and recombinant phages carrying tubulin genes were isolated by screening the library with heterologous chicken tubulin cDNA probes. The results of restriction endonuclease and hybridization analysis of DNA isolated from recombinant phages, and subcloned fragments thereof, were compatible with the restriction maps derived from digestion and Southern blot hybridization of genomic DNA.
TL;DR: Evidence is presented indicating that the protein antigens encoded by plasmids pLVS3 and pL VS5 are specific for pathogenic treponemal species.
Abstract: We have previously described the construction in Escherichia coli K-12 of a hybrid plasmid colony bank of Treponema pallidum (Nichols strain) genomic DNA. By screening a portion of this bank with an in situ immunoassay, we identified six E. coli clones that express T. pallidum antigens. In this study, the recombinant plasmids from each of these clones have been analyzed in E. coli maxicells and have been found to encode a number of proteins that are not of vector pBR322 origin and are, therefore, of treponemal origin. In each case, several of these proteins can be specifically precipitated from solubilized maxicell extracts by high-titer experimental rabbit syphilitic serum. Certain of these proteins are also precipitated by high-titer latent human syphilitic sera (HSS). The T. pallidum DNA inserts in these plasmids range in size from 6.2 to 14 kilobase pairs, and from the restriction patterns of the inserts and the protein profiles generated by each plasmid in maxicells, it is apparent that we have recovered a total of four unique clones from our colony bank. Recombinant plasmids pLVS3 and pLVS5 were of particular interest. Plasmid pLVS3 encodes three major protein antigens with molecular weights of 39,000, 35,000, and 25,000. These three proteins, which were not recognized by pooled normal human sera, were efficiently precipitated by most secondary HSS, latent HSS, and late HSS tested. These proteins were also precipitated, although somewhat inefficiently, by most primary HSS tested. Plasmid pLVS5 encodes a major protein antigen with a molecular weight of 32,000 and several minor protein antigens that, although efficiently precipitated by experimental rabbit syphilitic serum, were generally not recognized by the various HSS tested. Evidence is presented indicating that the protein antigens encoded by plasmids pLVS3 and pLVS5 are specific for pathogenic treponemal species. We have also demonstrated that immunoglobulin G antibodies directed against these protein antigens can be detected in rabbits experimentally infected with T. pallidum Nichols as early as 11 days postinfection.
TL;DR: The four ribosomal RNA genes of the rodent malaria parasite were identified and mapped by restriction enzyme analysis and Southern blot hybridization of genomic DNA and restriction analysis confirms both the low copy number and that these genes are not in an easily recognizable tandem array.
TL;DR: The data suggest that extensive introns were absent when restriction enzyme digests of DNA from cultures of paramecia with active and inactive genes were probed with portions of the cloned genes and no evidence for rearrangements or changes in gene dosage was found.
Abstract: Surface proteins from 11 antigenic types of Paramecium tetraurelia vary in molecular weight from 251,000 to 308,000. The size of a series of polyadenylated RNAs obtained from these types were correlated with the sizes of the proteins and judged to be the mRNAs for the proteins. The mRNAs were used to identify genomic DNA clones containing complementary sequences. The gene for antigen A was present in one copy per genome, and the data suggest that extensive introns were absent. When restriction enzyme digests of DNA from cultures of paramecia with active and inactive genes were probed with portions of the cloned genes, no evidence for rearrangements or changes in gene dosage was found.
Journal Article•
TL;DR: Using a series of human-Chinese hamster somatic cell hybrids containing unique combinations of human chromosomes, the alpha 1-antitrypsin gene as well as the sequence-related gene have been assigned to human chromosome 14 by Southern hybridization and synteny analysis.
Abstract: alpha 1-Antitrypsin is a major plasma protease inhibitor synthesized in the liver. Genetic deficiency of this protein predisposes the affected individuals to development of infantile liver cirrhosis or chronic obstructive pulmonary emphysema. The human chromosomal alpha 1-antitrypsin gene has been cloned and shown to contain three introns in the peptide-coding region. When the cloned alpha 1-antitrypsin gene was used as a hybridization probe to analyze Eco RI-digested genomic DNA from different individuals, two distinct bands of 9.6 kilobases (kb) and 8.5 kb in length were observed in every case. Further analysis using only labeled intronic DNA as the hybridization probe has indicated that the authentic alpha 1-antitrypsin gene resides within the 9.6-kb fragment. Thus the 8.5-kb fragment must contain another gene that is closely related in sequence to the alpha 1-antitrypsin gene. Using a series of human-Chinese hamster somatic cell hybrids containing unique combinations of human chromosomes, the alpha 1-antitrypsin gene as well as the sequence-related gene have been assigned to human chromosome 14 by Southern hybridization and synteny analysis.