RESUMO
A plasmid which contains a cos site of lambda and can be packaged into lambda bacteriophage particles is termed a "cosmid". Such plasmids can be used as gene cloning vectors in conjunction with an in vitro packaging system. The properties of a new series of cosmids based on the ColE1 replicon are described, including small temperature-sensitive plasmids which have lost mobilisation functions and carry no IS sequences. Amongst these plasmids are vectors for XmaI, BglII, BamHI, HindIII, PstI, KpnI, SalI and EcoRI. It is demonstrated that by using cosmids in particular size ranges these plasmids provide a high efficiency cloning system which yields essentially only hybrid clones without resort to a second selection or screening step, and without prior modification (e.g. phosphatase) treatment of the DNA. Attempts were made to optimise the cloning properties of the cosmid system. An Escherichia coli "gene bank" was obtained with an efficiency of 5 . 10(5) clones per microgram of E. coli DNA, and in which any particular unselected marker may be found in about one out of every 400 clones. It was demonstrated that deletion of mobilisation functions leads to loss of ability to form relaxation-complex without affecting copy number or segregation properties of the temperature-sensitive derivatives. The vectors are amplifiable in chloramphenicol to make up about 50% of the total cellular DNA.
Assuntos
Colífagos/genética , DNA Recombinante/metabolismo , Escherichia coli/genética , Plasmídeos , Colífagos/efeitos dos fármacos , Colífagos/metabolismo , Resistência Microbiana a Medicamentos , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Genes Virais , Rifampina/farmacologia , Transdução Genética , Transformação Bacteriana/efeitos dos fármacos , Replicação ViralAssuntos
Infecções por Echovirus/diagnóstico , Falência Hepática Aguda/etiologia , Viremia/diagnóstico , Diagnóstico Diferencial , Intervalo Livre de Doença , Infecções por Echovirus/complicações , Infecções por Echovirus/terapia , Enterovirus Humano B/classificação , Enterovirus Humano B/isolamento & purificação , Evolução Fatal , Feminino , Humanos , Recém-Nascido , Falência Hepática Aguda/diagnóstico , Falência Hepática Aguda/terapia , Testes de Função Hepática , Masculino , Países Baixos , Sorotipagem , Viremia/complicações , Viremia/terapiaRESUMO
Penicillin acylase is processed from a 90-kD precursor through the cleavage of a leader peptide and two further endopeptidase cleavages to yield an enzyme that contains a 22-kD (or 23-kD) and a 65-kD subunit. The endopeptidase cleavages require an intact carboxy terminus. This type of processing appears to be unique for a prokaryotic enzyme, having its most closely related analog in the synthesis and processing of preproinsulin and other eukaryotic hormones.