Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A vaccine strain of fowlpox virus (FPV) was genetically engineered to produce avian influenza virus hemagglutinin (HA). This was accomplished by inserting a cDNA copy of the avian influenza virus HA gene, which was regulated by a vaccinia virus promoter, into the FPV thymidine kinase (TK) gene. Two types of recombinant viruses, differing only in the orientation of the HA gene relative to an adjacent foreign gene (lacZ), were created. Following preliminary identification of FPV recombinants based on the generation of beta-galactosidase (lacZ gene product), correct insertion of the HA gene into the genomes of these viruses was verified by hybridization studies. Susceptible chickens vaccinated with these FPV recombinants produced specific hemagglutination-inhibiting antibodies against the HA antigen. In view of this immune response, these viruses may serve as vaccines against avian influenza virus. In this regard, they appeared to be less virulent than the parental virus.
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PMID:Expression of avian influenza virus hemagglutinin by recombinant fowlpox virus. 185 14

About 100 nucleotides of DNA sequence at the 5' noncoding region of the Choristoneura biennis entomopoxvirus spheroidin gene was chemically synthesized and inserted into a vaccinia expression vector, interrupting the vaccinia thymidine kinase gene. When the bacterial beta-galactosidase gene was introduced downstream of this sequence and a recombinant vaccinia virus containing these inserts was obtained by homologous recombination, beta-galactosidase was shown to be expressed at a high level late in the vaccinia infection cycle. The level of beta-galactosidase expression was four- to fivefold higher with this spheroidin-vaccinia recombinant virus than with a similar recombinant in which the beta-galactosidase gene was under the control of the vaccinia 7.5-kDa promoter. Primer extension and S1 mapping of the 5' terminus of the beta-galactosidase transcript located the transcription initiation site within the spheroidin DNA sequence, confirming the promoter nature of this DNA sequence in the vaccinia system. Dot blot analysis indicated that the difference in beta-galactosidase expression with these two recombinant viruses can be attributed to the difference in their transcript levels. We also demonstrated that full promoter activity encoded in the spheroidin 5' noncoding sequence was contained within a 38-nucleotide DNA fragment.
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PMID:The 5' noncoding region sequence of the Choristoneura biennis entomopoxvirus spheroidin gene functions as an efficient late promoter in the mammalian vaccinia expression system. 189 70

The transient expression of the two reporter genes, the genes for luciferase and bacterial beta-galactosidase, were used for comparative estimation of vaccinia viral promoters and for characterizing of the constructed plasmids. The recombinant clones of vaccinia virus expressing simultaneously and with high efficiency the luciferase and beta-galactosidase were used for studying the reproduction of vaccinia virus in mammalian cells. The advantages of the luciferase gene in using it as a reporter gene are discussed.
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PMID:[Expression of the glow worm luciferase gene in mammalian cells using vectors based on vaccinia viruses]. 190 77

Two genetics markers: the herpes simplex virus thymidine kinase and Escherichia coli beta-galactosidase genes were inserted into the 36K protein gene of vaccinia virus located in a HindIII-P DNA fragment. An unstability of recombinant viruses with Lac(+)-phenotype were discovered. A mechanism of viruses unstable variants formation was proposed, it was confirmed by the results of hybridisation analyses of virus recombinant genomes. The importance of a late nonstructural 36K protein gene for virus reproduction was demonstrated.
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PMID:[Molecular biological study of the vaccinia virus genome. IV. The late nonstructural 36K protein of vaccinia virus is vitally important]. 190 41

Using a recombinant vaccinia virus vector, the fiber protein from adenovirus serotype 2 has been expressed in human cells; the protein expressed was correctly assembled into trimers, glycosylated, and transported to the nucleus. Deletion of amino acids 2-5 (KRAR) resulted in accumulation of fiber in the cytoplasm; fusion of the sequence TKRVRL, found at the beginning of Ad7 fiber, to the N-terminus of this mutant restored correct targeting. Changing the charge of amino acids 91 and 92 within another potential targeting sequence (LKKTK to LEETK) had little effect on nuclear targeting. When fused to the N-terminus of beta-galactosidase and expressed in recombinant vaccinia virus, neither MKRARP nor MTKRVRL (from Ad2 and Ad7 fibers, respectively), were sufficient for efficient transport of the hybrid protein to the nucleus; on the other hand, fusions of either MKRARPSEDTF (from Ad2 fiber) or of MKRPRP (a known targeting sequence from the C-terminus of Ad2 E1A proteins) to beta-galactosidase were localized to the nucleus. These results suggest that sequences at the N-terminus of Ad2 and Ad7 fiber are required for correct nuclear targeting.
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PMID:The amino terminus of the adenovirus fiber protein encodes the nuclear localization signal. 196 47

We constructed a plasmid that contains a small piece of DNA with two vaccinia promoters running in opposite directions--a promoter from a late gene encoding an 11 K polypeptide (P11) and a promoter from an early gene encoding 25K (P25). These promoters were isolated from the Tian Tan strain of vaccinia virus and were flanked by the thymidine kinase (TK) sequence of the same virus. Genes encoding the hepatitis B virus surface antigen (HBsAg) and the Escherichia coli beta-galactosidase (LacZ) were inserted downstream of the 11 K and 25 K promoters respectively so that coexpression plasmids were constructed. Recombinant vaccinia viruses were selected directly by picking blue plaques formed under overlaying agarose medium containing X-gal. HBsAg was expressed to high level by these recombinant viruses. These recombinant viruses showed reduced virulence on rabbit skin and induced anti-HBs after intradermal inoculation of rabbits.
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PMID:Selection of recombinant vaccinia viruses (Tian Tan strain) expressing hepatitis B virus surface antigen by using beta-galactosidase as a marker. 211 Nov 42

A DNA fragment from fowlpox virus cloned on a plasmid vector was modified to contain foreign DNA inserts within an intergenic region. In a first step, a 32-base-pair intergenic region from the fowlpox virus genome corresponding to the position of the thymidine kinase locus in the vaccinia virus genome was enlarged to 55 base pairs by site-directed mutagenesis. A unique restriction endonuclease site introduced upstream of the intergenic region was then used to insert various foreign DNA fragments. The lacZ gene encoding beta-galactosidase and the measles virus gene encoding the fusion protein were positioned downstream of two vaccinia virus p7.5 promoter elements in either a direct repeat or inverted repeat orientation. Foreign DNA inserts contained within the fowlpox virus sequence were transferred to the viral genome by homologous recombination occurring in cells infected with a fowlpox virus temperature-sensitive mutant and transfected with both wild-type viral DNA and plasmid DNA. Recombinant viruses were selected for the expression of beta-galactosidase activity by screening for blue plaques in the presence of a chromogenic substrate. Stable recombinants expressing both the lacZ gene and the unselected measles gene were obtained when the p7.5 promoter was present as an inverted repeat. However, when the p7.5 promoter was in the direct repeat orientation, viral recombinants which initially expressed both gene inserts readily deleted the lacZ gene flanked by the promoter repeat. The methods described enable precise insertion and deletion of foreign genes in the fowlpox virus genome and could be applied to other intergenic regions of the same virus as well as other poxviruses.
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PMID:Construction of fowlpox virus vectors with intergenic insertions: expression of the beta-galactosidase gene and the measles virus fusion gene. 215 22

A transient expression assay for fowlpox virus (FPV) was developed to assess the feasibility of using heterologous promoters in FPV and to qualitatively determine relative promoter strength. A transient expression system for FPV has not been reported, and various methods used for transient expression in vaccinia-virus-infected cells produced negative results when used with FPV. Here a successful method for transient expression of E. coli beta-galactosidase in FPV-infected chick embryo fibroblasts is reported. This transient expression assay has been developed to qualitatively assess promoter recognition and gene expression by FPV. It should also prove useful in the identification of promoters from the FPV genomic library and in testing the accuracy of chimeric promoter-gene constructs.
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PMID:Transient expression assay for qualitative assessment of gene expression by fowlpox virus. 216 Nov 57

A quantitative and qualitative comparison of vaccinia virus (VV) promoter activity in fowlpox virus (FPV) and VV recombinants was performed. The VV PL11 late promoter was used to express beta-galactosidase from the E. coli LacZ gene in FPV (FPV-LacZ) and VV (VV-LacZ) recombinants. Time courses of FPV-LacZ beta-galactosidase expression in chicken embryo skin (CES) cells demonstrated temporal regulation of the PL11 promoter with maximum enzyme activity nine- and four-fold lower than those obtained in VV-LacZ infected 143B and CES cells, respectively. The level of beta-galactosidase activity per LacZ DNA gene copy was determined for each recombinant and found to be greater for VV-LacZ than FPV-LacZ. The VV P7.5 early/late promoter was used to express the E. coli xanthine-guanine phosphoribosyl transferase (Ecogpt) gene in FPV and VV recombinants. Northern blot analysis showed early Ecogpt RNA transcripts to be of defined lengths. Transcript size estimations mapped the termination sites to regions containing sequences associated with VV early transcript termination, providing supportive evidence for a common poxvirus early transcript termination signal. Late LacZ and Ecogpt transcripts were heterogeneous in length. S1 nuclease mapping of the 5'-ends of early and late Ecogpt RNA transcripts produced by FPV and VV recombinants showed transcription initiation occurred at the same sites in both poxviruses and corresponded to the regions previously identified as the early and late start sites of the P7.5 promoter. These results would indicate a high level of conservation in the expression and regulation of genes by poxviruses.
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PMID:Comparative analysis of vaccinia virus promoter activity in fowlpox and vaccinia virus recombinants. 216 93

Characterization of a late promoter of fowlpox virus (FPV) and a study of its activity in FPV and vaccinia virus (VV) was carried out. The 5'-mRNA start site of the FPV late gene mapped to a TAAAT sequence near the translation start site (ATG). A cloned DNA fragment of FPV genome (PFL1) comprising of the 5'-end of the late gene was used to express the LacZ gene of E. coli in FPV and VV recombinants. A comparative analysis of beta-galactosidase (BG) expression from the LacZ gene under the control of the FPV promoter and a VV late promoter (PL11) was performed. Like FPV-PL11-LacZ and VV-PL11-LacZ constructs, FPV-PFL1-LacZ and VV-PFL1-LacZ virus recombinants expressed BG indicating that essential features of transcription were conserved in the two viruses. Furthermore, the LacZ transcripts originating from PFL1 in FPV and VV recombinants mapped to the expected TAAAT sequence. Time course analysis of BG expressed by VV and FPV recombinants suggested that although the transcription machinery in the two viruses was essentially conserved, subtle differences in the efficiency of transcription or translation may exist.
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PMID:Activity of a fowlpox virus late gene promoter in vaccinia and fowlpox virus recombinants. 216 65


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