Bacteriophage

Abstract: Antibody fragments of predetermined binding specificity have recently been constructed from repertoires of antibody V genes, bypassing hybridoma technology and even immunization. The V gene repertoires are harvested from populations of lymphocytes, or assembled in vitro, and cloned for display of associated heavy and light chain variable domains on the surface of filamentous bacteriophage. Rare phage are selected from the repertoire by binding to antigen; soluble antibody fragments are expressed from infected bacteria; and the affinity of binding of selected antibodies is improved by mutation. The process mimics immune selection, and antibodies with many different binding specificities have been isolated from the same phage repertoire. Thus human antibody fragments have been isolated with specificities against both foreign and self antigens, including haptens, carbohydrates, secreted and cell surface proteins, viral coat proteins, and intracellular antigens from the lumen of the endoplasmic reticulum and ...

Abstract: Part 1 Growth maintenance and general techniques, C.R.Harwood and A.R.Archibald: the genus Bacillus growth of Bacillus maintenance and shipping of strains cell permeabilization and breakage radiolabelling of macromolecules. Part 2 Genetic analysis, S.M.Cutting and P.B.Vander Horn: mutagenesis transformation PBSI generalized transduction protoplast techniques selection and screening of recombinants genetic mapping complementation analysis. Part 3 Plasmids, S.Bron: plasmids used in Bacillus isolation of plasmid DNA plasmid DNA transfer systems for Bacillus plasmid replication in Bacillus plasmid instability in Bacillus subtilis efficient cloning systems for Bacillus subtilis based on pTA1060 and pAMbeta1. Part 4 Gene cloning techniques, J.Errington: cloning vectors cloning systems special purpose cloning vectors special purpose hosts. Part 5 Use of transposons and integrational vectors mutagenesis and construction of gene fusionson Bacillus species, P.Youngman: insertional mutagenesis with Tn917 insertional mutagenesis with integrational vectors using transposons or integrational vectors to tag specific regions of the chromosome cloning of chromosomal sequences adjacent to transposon insertions or plasmid integrations construction of lacZ fusions using transposons, integrational vectors and temperate phages information concerning vectors, strains and protocols. Part 6 Measuring gene expression in Bacillus, C.P.Moran: transcription products translation products transcription in vitro. Part 7 DNA repair and replication in Bacillus, R.E.Yasbin et al: DNA repair mechanisms DNA repair in Bacillus DNA replication. Part 8 The Bacillus cell envelope and secretion, C.R.Harwood et al: structure and function of the cell envelope synthesis and maturation of the Bacillus cell wall protein secretion electron microscopy for bacterial cells electron microscope methodologies. Part 9 Sporulation, W.L.Nicholson and P.Setlow: induction of sporulation sporulation - specific marker events isolation and analysis of small, acid-soluble spore proteins extraction and analysis of spore coat proteins sequential gene expression compartmentalization purification of spores storage of purified spores isolation and characterization of germination mutants isolation of spore growth mutants isolation of sporulation mutants. Part 10 Bacteriophages, H.E.Hemphill: host bacteria for growing B. subtilis phages media for culturing and maintaining B. subtilis phages culturing and maintaining bacteriophage stocks titering B. subtilis phages batch culturing of bacteriophages lysogens of temperate B. subtilis bacteriophages phage isolation and purification isolation of bacteriophage bacteriophage mutants. (Part contents).