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Titolo:
ELECTROTRANSFORMATION OF STREPTOCOCCUS-AGALACTINE WITH PLASMID DNA
Autore:
RICCI ML; MANGANELLI R; BERNERI C; OREFICI G; POZZI G;
Indirizzi:
IST SUPER SANITA,VIALE REGINA ELENA 299 I-00161 ROME ITALY IST SUPER SANITA I-00161 ROME ITALY UNIV SIENA,DIPARTIMENTO BIOL MOLEC,SEZ MICROBIOL I-53100 SIENA ITALY IST ZOOPROFILATTICO SPERIMENTALE LOMBARDO & EMILI I-25100 BRESCIA ITALY
Titolo Testata:
FEMS microbiology letters
fascicolo: 1-2, volume: 119, anno: 1994,
pagine: 47 - 52
SICI:
0378-1097(1994)119:1-2<47:EOSWPD>2.0.ZU;2-A
Fonte:
ISI
Lingua:
ENG
Soggetto:
GROUP-B STREPTOCOCCUS; CAPSULAR POLYSACCHARIDE ANTIGEN; GENETIC-TRANSFORMATION; ESCHERICHIA-COLI; STRUCTURAL DETERMINATION; ENTEROCOCCUS-FAECALIS; LACTOBACILLUS-CASEI; ELECTROPORATION; EXPRESSION; LACTOCOCCUS;
Keywords:
STREPTOCOCCUS AGALACTIAE; ELECTROTRANSFORMATION; SHUTTLE VECTOR;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
34
Recensione:
Indirizzi per estratti:
Citazione:
M.L. Ricci et al., "ELECTROTRANSFORMATION OF STREPTOCOCCUS-AGALACTINE WITH PLASMID DNA", FEMS microbiology letters, 119(1-2), 1994, pp. 47-52

Abstract

A protocol for efficient electrotransformation of Streptococcus agalactiae (group B streptococcus) Lancefield's strain O90R (NTCT 9993) (anunencapsulated derivative of type Ia strain O90) was developed. The Escherichia coli-Streptococcus shuttle vector pDP28 (7.8 kb) carrying the ermB gene for resistance to erythromycin was used as donor DNA. Frozen 'electrocompetent' cells were prepared by repeated washes in 10% glycerol. A 50-mu l aliquot containing about 5 X 10(9) colony forming units of bacteria was subjected to the electric pulse. Optimal conditions for electrotransformation were determined using different media, harvesting cells at different points of the growth curve, and using different field strengths. The dose-response curve for transformation of S. agalactiae with pDP28 showed one-hit kinetics as donor DNA varied between 0.01 and 3 mu g. The efficiency of electrotransformation for this range of amounts of donor DNA was 1.2 x 10(4) cfu mu g(-1). The transformation frequencies obtained with this electroporation protocol arehigh enough to allow both subcloning and shotgun cloning of streptococcal DNA in S. agalactiae.

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Documento generato il 24/11/20 alle ore 06:58:26