Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Gene Library”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 523 records · Page 29Linked to original sources
PubMed1994

Genes encoding homologues of three consecutive enzymes in the butyrate/butanol-producing pathway of Clostridium acetobutylicum are clustered on the Clostridium difficile chromosome.

Screening of a Clostridium difficile lambda EMBL3 gene library with antisera raised against C. difficile culture supernatant identified several clones expressing a 31-kDa protein. A 1.8-kb HindIII fragment subcloned from one of the clones was sufficient for expression of the 31-kDa polypeptide. Southern blot analysis showed a region homologous to this fragment to be present in all of 13 different C. difficile strains tested. Sequence analysis of the 1.8-kb fragment revealed three adjacent open reading frames. A database search showed that these three open reading frames appeared to encode homologues of three consecutive enzymes in the butanol/butyrate-producing pathway of Clostridium acetobutylicum (crotonase, beta-hydroxybutyryl coenzyme A dehydrogenase and thiolase).

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1986

Isolation of the DNA sequence coding indole-3-glycerol phosphate synthetase and phosphoribosylanthranilate isomerase of Schizophyllum commune.

A Schizophyllum gene library was made in plasmid pRK9. Plasmids from this library were tested for their ability to complement several auxotrophic mutations of Escherichia coli. The goal was to isolate a Schizophyllum auxotrophic gene that could be used to transform a corresponding Schizophyllum auxotrophic mutant to prototrophy. Complementation was observed only for E. coli trpC indole 3-glycerol phosphate synthetase (IGPS) and phosphoribosyl-anthranilate isomerase (PRAI) mutations. Plasmids with a Schizophyllum sequence coding for both IGPS and PRAI activities were recovered from E. coli transformants. Expression of the Schizophyllum gene (TRP1) in E. coli is probably dependent on the Serratia marcescens promoter of plasmid pRK9. The DNA sequence containing the Schizophyllum TRP1 gene was not obviously rearranged in cloning.

Aldose-Ketose Isomerases↗
PubMed1984

Nucleotide sequence of a yeast tRNAArg3A gene and its transcription in a homologous in vitro system.

Twelve bacterial clones containing complementary sequences to yeast tRNAArg3 were isolated from a gene library. The size of the yeast BamHI inserts ranges from 5.4 to 10 MDa. There are at least 6 copies of this gene in different loci of the yeast genome. Insert from clone pYAT-3 was mapped, and the presence of a tRNAArg3A gene was confirmed by DNA sequence. The coding region is colinear with the transcriptional product. Unlike other reported tRNAArg3A genes, this one is not linked to a tRNAAsp gene. In vitro transcription using a yeast extract produces a transcript of 76 +/- 1 bases.

Base Sequence↗
PubMed1982

A new approach for molecular cloning in cyanobacteria: cloning of an Anacystis nidulans met gene using a Tn901-induced mutant.

A new strategy for molecular cloning in the cyanobacterium Anacystis nidulans R-2 is described. This strategy involved the use of a transposon and was developed for the cloning of a gene encoding methionine biosynthesis. A met::Tn901 mutant was isolated. Chromosomal DNA fragments were cloned in the Escherichia coli plasmid vector pACYC184. A recombinant plasmid carrying the inactivated met::Tn901 gene was selected after transformation to E. coli. The cloned met::Tn901 DNA fragment was used as a probe to select the corresponding A. nidulans R-2 wild-type met gene from a gene library prepared in E. coli, using the newly constructed shuttle cosmid vector pPUC29. When transformed into A. nidulans Met- mutants, this cloned gene allowed the mutants to grow prototrophically.

Cloning, Molecular↗
PubMed2000

Nucleotide sequencing and transcriptional analysis of two tandem genes encoding glucosyltransferase (water-soluble-glucan synthetase) in Streptococcus cricetus HS-6.

Two tandem genes encoding glucosyltransferase synthesizing water-soluble glucan (GTF-S) were cloned from the lambda gene library of Streptococcus cricetus HS-6 (serotype a) using anti-GTF-S antibody, and the nucleotide sequences were analyzed. The two genes (ORF1 and ORF2) were identified as streptococcal glucosyltransferases based on the following evidence: [1] the deduced amino acid sequences of their products have an active site for catalytic action and C-terminal repeated units for dextran binding, and [2] a homology search revealed that the ORF1 and ORF2 products are homologous to the GtfS protein (77.4%) of S. downei Mfe28 and GtfT protein (83.8%) of S. sobrinus OMZ176, respectively, which are both known to have GTF-S activity. Therefore, ORF1 and ORF2 might be designated gtfS and gtfT of S. cricetus, respectively. A Northern blotting and RNase protection assay suggested that the gtfS and gtfT of S. cricetus are transcribed as a single bicistronic mRNA as well as separate monocistronic mRNAs. Primer extension analysis indicated multiple transcriptional start points for each gene.

Amino Acid Sequence↗
PubMed1992

Isolation of arginine auxotrophs, cloning by mutant complementation, and sequence analysis of the argC gene from the cyanobacterium Anabaena species PCC 7120.

Arginine auxotrophs of the dinitrogen-fixing cyanobacterium Anabaena species strain PCC 7120 were isolated after ultraviolet light mutagenesis and penicillin enrichment. Two of these auxotrophs were complemented by a cosmid gene library of the wild-type strain established in Escherichia coli that was transferred en masse to the mutants by conjugation. The gene complementing one of those mutants was found to complement an E. coli argC mutant. Sequencing analysis of the gene showed that it encodes a 322-residue polypeptide that is homologous to the ArgC protein of E. coli, Bacillus subtilis and Streptomyces clavuligerus and to the C-terminal moiety of the Saccharomyces cerevisiae ARG5,6 gene product, N-acetylglutamate semialdehyde dehydrogenase. A cysteine residue present in a highly conserved domain in the five proteins is probably located in the active site of the enzyme. Conserved among the ArgC proteins, sequences resembling the primary structure of nucleotide-binding domains are also found. Downstream of the Anabaena argC gene seven nearly perfect repeats of a heptanucleotide (consensus sequence:5'-CTAATGA-3') are found.

Aldehyde Oxidoreductases↗
PubMed1998

Cloning of GAP Gene from K. fragilis.

The K. fragilis CBS 397 gene library was screened with a GAP probe, which was designed according to the homology with S. cerevisiae, K. lactis, K. marxianus GAP gene. One positive clone pG1 containing GAP1 gene was isolated and confirmed by Southern hybridization. The GAP1 gene was partially sequenced. By using a fragment of the clone as a probe, another positive clone pG2 was acquired and also confirmed by Southern hybridization. The GAP2 gene from pG2 was completely sequenced. The upstream sequences of both genes were shown to have promoter activity. The fragment of pG1 could hybridize with three chromosomes of K. fragilis.

Journal Article↗
PubMed1993

Utilization of the mouse large intestine to select an Escherichia coli F-18 DNA sequence that enhances colonizing ability and stimulates synthesis of type 1 fimbriae.

Escherichia coli F-18, a normal human fecal isolate, is an excellent colonizer of the streptomycin-treated mouse large intestine. E. coli F-18 Col-, a derivative of E. coli F-18 which no longer makes the E. coli F-18 colicin, colonizes the large intestine as well as E. coli F-18 when fed to mice alone but is eliminated when fed together with E. coli F-18. Random sequences of E. coli F-18 DNA were cloned into pRLB2, a par-B-stabilized derivative of pHC79. The entire gene library was transformed into E. coli F-18 Col- and fed to streptomycin-treated mice. The mouse large intestine selected a predominant clone which contained a recombinant plasmid (pRLB7) that enhanced E. coli F-18 Col- colonizing ability 100-fold but did not stimulate colicin synthesis. Moreover, pRLB7 simultaneously improved the survival of E. coli F-18 Col- in stationary phase in vitro, utilizing nutrients derived from mouse cecal mucus, and stimulated synthesis of both type 1 fimbriae and three E. coli F-18 Col- outer membrane proteins (74, 71, and 69 kDa). The 6.5-kb E. coli F-18 DNA sequence in pRLB7 does not contain either the fim operon or pilG (hns), both known to be involved in type 1 fimbrial synthesis. The sequence encodes six proteins, all smaller than the three E. coli F-18 Col- outer membrane proteins whose synthesis it stimulates. Collectively, the results suggest that the cloned E. coli F-18 DNA sequence contains one or more regulators of E. coli F-18 Col- operons expressed in the mouse large intestine in vivo and in isolated mouse cecal mucus in vitro.

Animals↗
PubMed2004

Design of a gene family screening library targeting G-protein coupled receptors.

An iterative process for the design of a G-protein coupled receptor (GPCR) gene family screening library has been developed. A key element of this process is the computational generation of pharmacophore descriptors of known GPCR ligands. Subsequent iterative analysis allows prioritization of scaffolds and sub-libraries for inclusion in the library. The final library, which consisted of 13,769 compounds, displayed a 2.6% hit rate when screened against the micro-opioid receptor.

Combinatorial Chemistry Techniques↗
PubMed1995

Molecular cloning of Rab-related genes in the yeast Yarrowia lipolytica. Analysis of RYL1, an essential gene encoding a SEC4 homologue.

Small GTP-binding proteins of the Rab family are involved in the vesicular traffic inside eukaryotic cells. A gene library from the yeast Yarrowia lipolytica was screened with an oligonucleotide deduced from a highly conserved sequence in the Rab family. Four different genes were isolated. One of them, RYL1, was shown to be essential for cell viability. RYL1p displayed a high similarity with and tight phylogenetic relationships to SEC4p. When placed under the control of the GAL10 promoter, RYL1 was able to specifically relieve the thermosensitivity of a sec4-8 mutant of Saccharomyces cerevisiae. Therefore, it is proposed that RYL1 is a functional homologue of the S. cerevisiae SEC4 gene and is involved in the fusion of secretory vesicles with the plasma membrane in the general protein secretion pathway.

Amino Acid Sequence↗
PubMed1982

Structure of human immunoglobulin gamma genes: implications for evolution of a gene family.

We have cloned five human immunoglobulin gamma genes from a fetal liver gene library. Four of them encode the known human immunoglobulin gamma chains gamma 1, gamma 2, gamma 3 and gamma 4. A fifth gamma gene seems to be a pseudogene. Nucleotide sequence determination demonstrates that the gamma 3 gene contains four separate hinge exons. Comparison of these hinge exons with those of the other gamma genes indicates that the first hinge exon is homologous to that of the pseudogene, and that the other three hinge exons are homologous to that of the gamma 1 gene, suggesting that the gamma 3 gene ancestor is a hybrid gene created by unequal crossing-over between the ancestral gamma 1 and psi gamma genes. Amplification of the gamma 1-type hinge exon probably followed to complete the gamma 3 gene. This hypothesis inevitably postulates the gene order 5'-gamma 1-gamma 3-psi gamma-3'. Cloning of overlapping chromosomal segments demonstrates that the gamma 2 gene is located 19 kb 5' to the gamma 4 gene. These analyses indicate that the human gamma-gene family has evolved by several types of DNA rearrangemet, including duplication of a complete gene; duplication of a hinge exon; and reassortment of exons by unequal cross-over between two adjacent genes.

Animals↗
PubMed2005

rdlA, a new gene encoding a rhodanese-like protein in Halanaerobium congolense and other thiosulfate-reducing anaerobes.

The recently described anaerobic moderately halophilic bacterium Halanaerobium congolense has been shown to reduce thiosulfate and sulfur-but not sulfate-into sulfide. When cultivated in the presence of thiosulfate as terminal electron acceptor, H. congolense possesses a highly active thiosulfate:cyanide sulfur-transferase activity (rhodanese-like enzyme). A gene library of H. congolense (DSM 11287T) was constructed, and a 3.1-kb Sau3A DNA that encompassed a thiosulfate:cyanide sulfur-transferase-encoding gene was isolated in Escherichia coli. This fragment contains 2 orfs, which were separately subcloned in E. coli. The 900-bp gene encoding the rhodanese-like protein was named rdlA. RdlA differs from other known rhodanese-like proteins by having two potential catalytic sites, one N-terminal and one C-terminal, both harboring a cysteine. The two putative active sites are preceded by a highly-conserved region of unknown function. Closely related genes were also characterized in other thiosulfate-reducing non-sulfate-reducing anaerobes belonging to phylogenetically distant microorganisms, thus suggesting that RdlA is of importance in the mechanism of thiosulfate reduction by numerous members of the domain Bacteria.

Amino Acid Sequence↗
PubMed1979

The ovalbumin gene: cloning and molecular organization of the entire natural gene.

We report the analyses of recently cloned restriction fragments of the natural ovalbumin gene that overlap in part with previously cloned DNA fragments but extend further into the flanking sequences of the gene. These clones now permit us to identify the DNA sequence that codes for the 5' end of ovalbumin mRNA. Based on these and previous results, the molecular organization of the entire ovalbumin gene was established. The entire gene is composed of eight structural DNA sequences separated by seven intervening sequences that are not present in the mature mRNA. In addition, an ovalbumin gene clone has been obtained from a chicken gene library. Analysis of DNA isolated from this particular clone by molecular hybridization and electron microscopic mapping revealed that it contains the entire ovalbumin gene a single segment of DNA and its structure was consistent with that predicted from our physical map constructed from individually cloned fragments of the gene.

Animals↗
PubMed1998

Isolation and characterization of EPD1, an essential gene for pseudohyphal growth of a dimorphic yeast, Candida maltosa.

Additional copies of the centromeric DNA (CEN) region induce pseudohyphal growth in a dimorphic yeast, Candida maltosa (T. Nakazawa, T. Motoyama, H. Horiuchi, A. Ohta, and M. Takagi, J. Bacteriol. 179:5030-5036, 1997). To understand the mechanism of this transition, we screened the gene library of C. maltosa for sequences which could suppress this morphological change. As a result, we isolated the 5' end of a new gene, EPD1 (for essential for pseudohyphal development), and then cloned the entire gene. The predicted amino acid sequence of Epdlp was highly homologous to those of Ggp1/Gas1/Cwh52p, a glycosylphosphatidylinositol-anchored protein of Saccharomyces cerevisiae, and Phr1p and Phr2p of Candida albicans. The expression of EPD1 was moderately regulated by environmental pH. A homozygous EPD1 null mutant showed some morphological defects and reduction in growth rate and reduced levels of both alkali-soluble and alkali-insoluble beta-glucans. Moreover, the mutant could not undergo the transition from yeast form to pseudohyphal form induced by additional copies of the CEN sequence at pH 4 or by n-hexadecane at pH 4 or pH 7, suggesting that EPD1 is not essential for yeast form growth but is essential for transition to the pseudohyphal form. Overexpression of the amino-terminal part of Epd1p under the control of the GAL promoter suppressed the pseudohyphal development induced by additional copies of the CEN sequence, whereas overexpression of the full-length EPD1 did not. This result and the initial isolation of the 5' end of EPD1 as a suppressor of the pseudohyphal growth induced by the CEN sequence suggest that the amino-terminal part of Epd1p may have a dominant-negative effect on the functions of Epd1p in the pseudohyphal growth induced by the CEN sequence.

Amino Acid Sequence↗
PubMed1997

Cloning and sequencing of draTG genes and their downstream region of Azospirillum brasilense Yu62.

An 8-kb fragment was cloned by probing the gene library of Azospirillum brasilense Yu62 with the 4.0-kb draTG fragment of A. brasilense Sp7. DNA hybridization of this fragment demonstrated that draTG genes were located in a 3.0-kb EcoR I-Kpn I fragment, and were contiguous to the nifH gene. This 3.0-kb fragment was completely sequenced on both strands. Sequence analysis of the fragment revealed that it included the full-length draTG genes and two ORFs downstream of draG (ORF3 and incomplete ORF4). The draTG and downstream ORF3 are presumed to be cotranscribed as a single operon. Promoter element analysis of the sequenced region showed that there were some elements of the sigma 54-dependent promoter (DPEs and UASs) in the upstream region of draG and ORF3. This suggests that the draG and ORF3 might be transcribed independently in addition to being cotranscribed with draT. Both the DNA and amino acid sequences of the draTG genes from A. brasilense Yu62 were compared with those of other nitrogen-fixing bacteria. The results showed the draTG genes were highly conservative. There were only a few changes among strains and/or species. Homology analysis of ORF revealed that the ORF3 immediately downstream of draG was homologous not only with the ORF at the same position in Rhodospirillum rubrum and A. lipoferum, but also with the ORF14 of Azotobacter vinelandii and the ORF4 showed extensive similarity to yafJ of Escherichia coli.

Amino Acid Sequence↗
PubMed1993

Cloning and sequence analysis of the X-prolyl-dipeptidyl-aminopeptidase gene (pepX) from Lactobacillus delbrückii ssp. lactis DSM7290.

Lactobacillus delbrückii ssp. lactis DSM7290 possesses an X-prolyl-dipeptidyl-aminopeptidase, designated PepX, which catalyses the hydrolytic removal of N-terminal dipeptidyl residues from peptides containing proline in the penultimate position. Using the specific substrate L-Ala-L-Pro-p-nitroanilide, PepX was purified by a four-step procedure including ammonium sulphate fractionation, hydrophobic interaction chromatography, ion exchange chromatography, and affinity chromatography. The N-terminus of the purified protein was sequenced. Screening of a gene library of chromosomal Lactobacillus delbrückii ssp. lactis DSM7290 DNA in the low-copy-number vector pLG339 resulted in the identification of the pepX gene in Escherichia coli using a specific plate assay with Gly-L-Pro-beta-naphthylamide as substrate. Nucleotide sequence analysis revealed an open reading frame of 2376 bp, coding for a protein of 792 amino acids with a molecular mass of 88449 Da.

Amino Acid Sequence↗
PubMed1989

The V(D)J recombination activating gene, RAG-1.

The RAG-1 (recombination activating gene-1) genomic locus, which activates V(D)J recombination when introduced into NIH 3T3 fibroblasts, was isolated by serial genomic transfections of oligonucleotide-tagged DNA. A genomic walk spanning 55 kb yielded a RAG-1 genomic probe that detects a single 6.6-7.0 kb mRNA species in transfectants and pre-B and pre-T cells. RAG-1 genomic and cDNA clones were biologically active when introduced into NIH 3T3 cells. Nucleotide sequencing of human and mouse RAG-1 cDNA clones predicts 119 kd proteins of 1043 and 1040 amino acids, respectively, with 90% sequence identity. RAG-1 has been conserved between species that carry out V(D)J recombination, and its pattern of expression correlates exactly with the pattern of expression of V(D)J recombinase activity. RAG-1 may activate V(D)J recombination indirectly, or it may encode the V(D)J recombinase itself.

Amino Acid Sequence↗
PubMed1981

Human ribosomal RNA gene spacer sequences are found interspersed elsewhere in the genome.

A cloned EcoRI fragment containing human 18 S rRNA gene sequences was used to screen a gene library to obtain a set of 8 overlapping cloned DNA segments extending into the non-transcribed spacer region of the human ribosomal RNA gene cluster. 19.4 kb of the approx. 43-kb rDNA repeat was obtained in cloned form and mapped with restriction endonucleases. None of the clones obtained extended into 28 S rRNA sequences. A 7-kb region of non-transcribed spacer DNA shared in common between five independent clones was subjected to comparative restriction digests. It was estimated that sequences among the five different spacer isolated varied by not more than 1.0%, if all the observed differences are assumed due to point mutation. HaeII-restriction fragments from within this same 7-kb region contain sequences carried not only within the tandem repeats of the gene cluster but interspersed elsewhere in the genome. Some of these sequences correspond to the Alu family of highly repeated interspersed sequences.

Base Sequence↗