Artificial regulation of gene expression in Escherichia coli by RNase P.

Plasmids encoding various external guide sequences (EGSs) were constructed and inserted into Escherichia coli. In strains harboring the appropriate plasmids, the expression of fully induced beta-galactosidase and alkaline phosphatase activity was reduced by more than 50%, while no reduction in such activity was observed in strains with non-specific EGSs. The inhibition of gene expression was virtually abolished at restrictive temperatures in strains that were temperature-sensitive for RNase P (EC 3.1.26.5). Northern blot analysis showed that the steady-state copy number of EGS RNAs was several hundred per cell in vivo. A plasmid that contained a gene for M1 RNA covalently linked to a specific EGS reduced the level of expression of a suppressor tRNA that was encoded by a separate plasmid. Similar methods can be used to regulate gene expression in E. coli and to mimic the properties of cold-sensitive mutants.

Mutagenesis of the human apolipoprotein B gene in a yeast artificial chromosome reveals the site of attachment for apolipoprotein(a).

Lipoprotein(a) [Lp(a)] is a lipoprotein formed by the disulfide linkage of apolipoprotein (apo) B100 of a low density lipoprotein particle to apolipoprotein(a). Prior studies have suggested that one of the C-terminal Cys residues of apo-B100 is involved in the disulfide linkage of apo-B100 to apo(a). To identify the apo-B100 Cys residue involved in the formation of Lp(a), we constructed a yeast artificial chromosome (YAC) spanning the human apo-B gene and used gene-targeting techniques to change Cys-4326 to Gly. The mutated YAC DNA was used to generate transgenic mice expressing the mutant human apo-B100 (Cys4326Gly). Unlike the wild-type human apo-B100, the mutant human apo-B100 completely lacked the ability to bind to apo(a) and form Lp(a). This study demonstrates that apo-B100 Cys-4326 is required for the assembly of Lp(a) and shows that gene targeting in YACs, followed by the generation of transgenic mice, is a useful approach for analyzing the structure of large proteins coded for by large genes.

Use of yeast artificial chromosomes (YACs) in studies of mammalian development: production of beta-globin locus YAC mice carrying human globin developmental mutants.

To test whether yeast artificial chromosomes (YACs) can be used in the investigation of mammalian development, we analyzed the phenotypes of transgenic mice carrying two types of beta-globin locus YAC developmental mutants: (i) mice carrying a G-->A transition at position -117 of the A gamma gene, which is responsible for the Greek A gamma form of hereditary persistence of fetal hemoglobin (HPFH), and (ii) beta-globin locus YAC transgenic lines carrying delta- and beta-globin gene deletions with 5' breakpoints similar to those of deletional HPFH and delta beta-thalassemia syndromes. The mice carrying the -117 A gamma G-->A mutation displayed a delayed gamma- to beta-globin gene switch and continued to express A gamma-globin chains in the adult stage of development as expected for carriers of Greek HPFH, indicating that the YAC/transgenic mouse system allows the analysis of the developmental role of cis-acting motifs. The analysis of mice carrying 3' deletions first provided evidence in support of the hypothesis that imported enhancers are responsible for the phenotypes of deletional HPFH and second indicated that autonomous silencing is the primary mechanism for turning off the gamma-globin genes in the adult. Collectively, our results suggest that transgenic mice carrying YAC mutations provide a useful model for the analysis of the control of gene expression during development.

Metaphase and interphase fluorescence in situ hybridization mapping of the rice genome with bacterial artificial chromosomes.

Fluorescence in situ hybridization (FISH) is a powerful tool for physical mapping in human and other mammalian species. However, application of the FISH technique has been limited in plant species, especially for mapping single- or low-copy DNA sequences, due to inconsistent signal production in plant chromosome preparations. Here we demonstrate that bacterial artificial chromosome (BAC) clones can be mapped readily on rice (Oryza sativa L.) chromosomes by FISH. Repetitive DNA sequences in BAC clones can be suppressed efficiently by using rice genomic DNA as a competitor in the hybridization mixture. BAC clones as small as 40 kb were successfully mapped. To demonstrate the application of the FISH technique in physical mapping of plant genomes, both anonymous BAC clones and clones closely linked to a rice bacterial blight-resistance locus, Xa21, were chosen for analysis. The physical location of Xa21 and the relationships among the linked clones were established, thus demonstrating the utility of FISH in plant genome analysis.

Assessment of Natural and Manmade Materials in US Artificial Reef Programs

Natural coral reefs are in a state of serious decline worldwide. The pressures of over fishing, recreational activities, environmental pollutants, and global warming have stressed these marine ecosystems to the breaking point. One of the oldest methods of augmenting natural reef systems is the implementation of artificial reefs. These projects are not as simple as dumping waste or scrap materials in offshore areas. Proper material selection is vital to produce a healthy artificial marine habitat that is completed on schedule and on budget. This Capstone Project will evaluate the most commonly used materials and provide a comparison of their strengths and weaknesses. This comparison provides a valuable tool for project managers as they begin the reef planning process.

The complete confectioner, or, Housekeeper's guide to a simple and speedy method of understanding the whole art of confectionary : the various ways of preserving and candying, dry and liquid, all kinds of fruit, nuts, flowers, herbs, &c. And the method of keeping them fresh and fine all the year round; the different ways of clarifying sugar; with directions for making fruit pastes, bomboons, pastils, compotes, fruit ices, cream ices, marmalades, jellies, jams, cakes, puffs, biscuits, tarts, custards, cheesecakes, sweetmeats, fritters, creams, syllabubs, blanc-mange, flummeries, ornaments for grand entertainments, dragees, syrups of all kinds, nicknacks and trifles for desserts, strong cordials, oils, simple waters, milk punch that will keep 20 years, and all sorts of English wines. Also, the art of making artificial fruit, with the stalks in it, so as to resemble the natural fruit. To which are added, some bills of fare for desserts for private families /

Originally published under title: The compleat confectioner. Cf. Bitting, K.G. Gastronomic bib., p. 190.

The ABC of carp culture. A treatise on the German, or European carp; its wonderful growth & fecundity, instructions for its propagation and culture as an article of food for sale; also minute directions for the construction of artificial lakes and ponds, comprising those from only a few yards in area to ponds and lakes covering acres /

Mode of access: Internet.

A system monitor for programs in artificial intelligence /

Bibliography: p. 84-85.