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At least 1,189 records · Page 66Linked to original sources
PubMed2000

Engineering the provitamin A (beta-carotene) biosynthetic pathway into (carotenoid-free) rice endosperm.

Rice (Oryza sativa), a major staple food, is usually milled to remove the oil-rich aleurone layer that turns rancid upon storage, especially in tropical areas. The remaining edible part of rice grains, the endosperm, lacks several essential nutrients, such as provitamin A. Thus, predominant rice consumption promotes vitamin A deficiency, a serious public health problem in at least 26 countries, including highly populated areas of Asia, Africa, and Latin America. Recombinant DNA technology was used to improve its nutritional value in this respect. A combination of transgenes enabled biosynthesis of provitamin A in the endosperm.

Alkyl and Aryl Transferases↗
PubMed1987

The tat gene of human T-lymphotropic virus type 1 induces mesenchymal tumors in transgenic mice.

Human T-lymphotropic virus type 1 (HTLV-1) is a suspected causative agent of adult T-cell leukemia. One of the viral genes encodes a protein (tat) that not only results in transactivation of viral gene expression but may also regulate the expression of certain cellular genes that are important for cell growth. Transgenic mice that expressed the authentic tat protein under the control of the HTLV-1 long terminal repeat were generated, and cell types that are permissive for the viral promoter and the effects of the tat gene on these cells were studied. Three of eight founder mice with high levels of expression of the transgene in muscle were bred and then analyzed. All developed soft tissue tumors at multiple sites between 13 to 17 weeks of age. This phenotype was transmitted to nine of nine offspring that inherited the tat gene and were available for analysis. The remaining five founders expressed the transgene in the thymus, as well as in muscle. This second group of mice all exhibited extensive thymic depletion and growth retardation; in all of these mice, death occurred between 3 to 6 weeks of age before tumors became macroscopically visible. The tat gene under the control of the HTLV-1 regulatory region showed tissue-specific expression and the tat protein efficiently induced mesenchymal tumors. The data establish tat as an oncogenic protein and HTLV-1 as a transforming virus.

Animals↗
PubMed2000

Mosquito engineering. Building a disease-fighting mosquito.

With resistance against insecticides on the rise and a U.N.-backed push to phase out DDT, several labs have embarked on the most ambitious and futuristic of all approaches to combat malaria: They hope to replace billions and billions of mosquitoes in the world's endemic areas with new strains, created in the lab, that would be "refractory," or unable to transmit Plasmodium, the parasite that causes the disease. The idea is not that farfetched, these researchers claim.

Aedes↗
PubMed1985

Constitutive and conditional suppression of exogenous and endogenous genes by anti-sense RNA.

Plasmid DNA directing transcription of the noncoding (anti-sense) DNA strand can specifically inhibit the expression of several test genes as well as normal, endogenous genes. The anti-sense plasmid constructions can be introduced into eukaryotic cells by transfection or microinjection and function in both transient and stable transformation assays. Anti-sense transcripts complementary to as little as 52 bases of 5' untranslated target gene mRNA specifically suppress gene activity as well as, or more efficiently than, anti-sense transcripts directed against the protein coding domain alone. Conditional anti-sense inhibition is accomplished with the use of hormone-inducible promoter sequences. Suppression of endogenous actin gene activity by anti-sense RNA is detected as a decrease in growth rate and as a reduction in the number of actin microfilament cables. These observations suggest that anti-sense RNA may be generally useful for suppressing the expression of specific genes in vivo and may be a potential molecular alternative to classical genetic analysis.

Acetyltransferases↗
PubMed1985

Amino acid replacements that compensate for a large polypeptide deletion in an enzyme.

Deletion of more than 400 amino acids from the carboxyl terminus of an enzyme causes a severe reduction in catalytic activity. Selected point mutations within the residual protein partially reverse the effects of the missing segment. The selection can yield mutants with activities at least ten times as high as those of the starting polypeptides. One well-characterized mutation, a single amino acid replacement in the residual polypeptide, increases the catalytic activity of the polypeptide by a factor of 5. The results suggest substantial potential for design of protein elements to compensate for missing polypeptide sequences. They also may reflect that progenitors of large aminoacyl-tRNA (transfer RNA) synthetases--one of which was used in these studies--were themselves much smaller.

Alanine-tRNA Ligase↗
PubMed1983

Prospects in plant genetic engineering.

The functional expression of a novel gene in a genetically engineered plant has not yet been reported. One major barrier in movement toward this goal is our limited understanding of the molecular bases of gene expression. Attempts to establish genetic engineering as a practical facet of plant breeding are also complicated by the fact that genes for most important plant characteristics have not yet been identified. However, the benefits to be gained from all aspects of plant improvement are stimulating research into both the development of plant transformation technology and the isolation and characterization of genes responsible for valuable traits. As scientists develop greater knowledge of plant molecular genetics, we can expect to see practical applications in such diverse areas as improvement of plant nutritional quality, decreases in fertilization requirements, and increases in resistance to environmental stresses and pathogens.

Agriculture↗
PubMed1994

Engineered biosynthesis of a complete macrolactone in a heterologous host.

Macrocyclic polyketides have been subjects of great interest in synthetic and biosynthetic chemistry because of their structural complexity and medicinal activities. With expression of the entire 6-deoxyerythronolide B synthase (DEBS) (10,283 amino acids) in a heterologous host, substantial quantities of 6-deoxyerythronolide B (6dEB), the aglycone of the macrolide antibiotic erythromycin, and 8,8a-deoxyoleandolide, a 14-membered lactone ring identical to 6dEB except for a methyl group side chain in place of an ethyl unit, were synthesized in Streptomyces coelicolor. The biosynthetic strategy utilizes a genetic approach that facilitates rapid structural manipulation of DEBS or other modular polyketide synthases (PKSs), including those found in actinomycetes with poorly developed genetic methods. From a technological viewpoint, this approach should allow the rational design of biosynthetic products and may eventually lead to the generation of diverse polyketide libraries by means of combinatorial cloning of naturally occurring and mutant PKS modules.

Acyl Coenzyme A↗
PubMed2006

Transformation of the green alga Haematococcus pluvialis with a phytoene desaturase for accelerated astaxanthin biosynthesis.

Astaxanthin is a high-value carotenoid which is used as a pigmentation source in fish aquaculture. Additionally, a beneficial role of astaxanthin as a food supplement for humans has been suggested. The unicellular alga Haematococcus pluvialis is a suitable biological source for astaxanthin production. In the context of the strong biotechnological relevance of H. pluvialis, we developed a genetic transformation protocol for metabolic engineering of this green alga. First, the gene coding for the carotenoid biosynthesis enzyme phytoene desaturase was isolated from H. pluvialis and modified by site-directed mutagenesis, changing the leucine codon at position 504 to an arginine codon. In an in vitro assay, the modified phytoene desaturase was still active in conversion of phytoene to zeta-carotene and exhibited 43-fold-higher resistance to the bleaching herbicide norflurazon. Upon biolistic transformation using the modified phytoene desaturase gene as a reporter and selection with norflurazon, integration into the nuclear genome of H. pluvialis and phytoene desaturase gene and protein expression were demonstrated by Southern, Northern, and Western blotting, respectively, in 11 transformants. Some of the transformants had a higher carotenoid content in the green state, which correlated with increased nonphotochemical quenching. This measurement of chlorophyll fluorescence can be used as a screening procedure for stable transformants. Stress induction of astaxanthin biosynthesis by high light showed that there was accelerated accumulation of astaxanthin in one of the transformants compared to the accumulation in the wild type. Our results strongly indicate that the modified phytoene desaturase gene is a useful tool for genetic engineering of carotenoid biosynthesis in H. pluvialis.

Biotechnology↗
PubMed1992

Insecticidal properties of genetically engineered baculoviruses expressing an insect juvenile hormone esterase gene.

Exploring the possibility of enhancing the properties of baculoviruses as biological control agents of insect pests, we tested the effect of expressing an insect gene (jhe) encoding juvenile hormone esterase. Juvenile hormone esterase inactivates juvenile hormone, which regulates the outcome of an insect molt. A cDNA encoding the juvenile hormone esterase of Heliothis virescens was inserted into the genome of Autographa californica nuclear polyhedrosis virus such that the gene was expressed under the control of a strong, modified viral promoter. This virus, however, naturally encodes an ecdysteroid UDP-glucosyltransferase which inactivates ecdysone, the hormone which initiates molting. Since ecdysteroid UDP-glucosyltransferase could mask the effects of jhe expression by blocking molting entirely, jhe-expressing viruses in which the ecdysteroid UDP-glucosyltransferase gene was deleted or disrupted were constructed. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of proteins from infected cells revealed several intracellular proteins and two major secreted proteins which reacted with antibodies to authentic juvenile hormone esterase. Western blot analysis coupled with tunicamycin treatment indicated that differential glycosylation was responsible for the multiple products. Hemolymph of recombinant virus-infected fourth-instar Trichoplusia ni larvae contained levels of juvenile hormone esterase activity 40-fold higher than maximal levels found in uninfected larvae. However, little or no difference in developmental characteristics, weight gain, or time of mortality was observed between insects infected with the jhe-expressing viruses and control viruses.

Animals↗
PubMed1992

Genetic analysis of a locus on the Bacteroides ovatus chromosome which contains xylan utilization genes.

Bacteroides ovatus, a gram-negative obligate anaerobe found in the human colon, can utilize xylan as a sole source of carbohydrate. Previously, a 3.8-kbp segment of B. ovatus chromosomal DNA, which contained genes encoding a xylanase (xylI) and a bifunctional xylosidase-arabinosidase (xsa), was cloned, and expression of the two genes was studied in Escherichia coli (T. Whitehead and R. Hespell, J. Bacteriol. 172:2408-2412, 1990). In the present study, we have used segments of the cloned region to construct insertional disruptions in the B. ovatus chromosomal locus containing these two genes. Analysis of these insertional mutants demonstrated that (i) xylI and xsa are probably part of the same operon, with xylI upstream of xsa, (ii) the true B. ovatus promoter was not cloned on the 3.5-kbp DNA fragment which expressed xylanase and xylosidase in E. coli, (iii) there is at least one gene upstream of xylI which could encode an arabinosidase, and (iv) xylosidase rather than xylanase may be a rate-limiting step in xylan utilization. Insertional mutations in the xylI-xsa locus reduced the rate of growth on xylan, but the concentration of residual sugars at the end of growth was the same as that with the wild type. Thus, a slower rate of growth on xylan was not accompanied by less extensive digestion of xylan. Mutants in which xylI had been disrupted still expressed some xylanase activity. This second activity was associated with membranes and produced xylose from xylan, whereas the xylI gene product partitioned primarily with the soluble fraction and produced xylobiose from xylan.

Bacteroides↗