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Titolo:
Autocloning and amplification of LIP2 in Yarrowia lipolytica
Autore:
Pignede, G; Wang, HJ; Fudalej, F; Seman, M; Gaillardin, C; Nicaud, JM;
Indirizzi:
INRA, Lab Microbiol & Genet Mol, Ctr Grignon, F-78850 Thiverval Grignon, France INRA Thiverval Grignon France F-78850 F-78850 Thiverval Grignon, France Lab Mayoly Spindler, Serv Rech, F-78401 Chatou, France Lab Mayoly Spindler Chatou France F-78401 v Rech, F-78401 Chatou, France
Titolo Testata:
APPLIED AND ENVIRONMENTAL MICROBIOLOGY
fascicolo: 8, volume: 66, anno: 2000,
pagine: 3283 - 3289
SICI:
0099-2240(200008)66:8<3283:AAAOLI>2.0.ZU;2-E
Fonte:
ISI
Lingua:
ENG
Soggetto:
COPY-NUMBER INTEGRATION; HIGH-LEVEL EXPRESSION; YEAST CANDIDA-UTILIS; RIBOSOMAL DNA; SACCHAROMYCES-CEREVISIAE; SCHIZOSACCHAROMYCES-POMBE; KLUYVEROMYCES-LACTIS; HANSENULA-POLYMORPHA; PROTEIN-PRODUCTION; MICROBIAL LIPASES;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Agriculture,Biology & Environmental Sciences
Life Sciences
Citazioni:
48
Recensione:
Indirizzi per estratti:
Indirizzo: Nicaud, JM INRA, Lab Microbiol & Genet Mol, Ctr Grignon, BP 01, F-78850 Thiverval Grignon, France INRA BP 01 Thiverval Grignon France F-78850 al Grignon, France
Citazione:
G. Pignede et al., "Autocloning and amplification of LIP2 in Yarrowia lipolytica", APPL ENVIR, 66(8), 2000, pp. 3283-3289

Abstract

We synthesized a Yarrowia lipolytica strain overproducing lipase for industrial applications by using long terminal repeat (zeta) of the Y. lipolytica retrotransposon Ylt1 and an allele of URA3 with a promoter deletion to construct JMP3. JMP3 is a derivative of plasmid pHSS6 carrying a NotI-NotI cassette which contains a defective URA3 allele, a polylinker sequence, and the zeta region for targeting to multiple sites in the genome of the recipient. We inserted the LIP2 gene (encoding extracellular lipase) under the control of the strong POX2 promoter into JMP3 to generate JMP6. The pHSS6 region was removed by NotI digestion prior to transformation. Two I: lipolyticastrains transformed with the JMP6 LIP2 cassette had a mean of 10 integrated copies devoid of the Escherichia coli region, corresponding to an autocloning event. The copy number in the transformants was stable even after 120 generations in nonselective and lipase-inducing conditions. The resulting strains could produce 0.5 g of active lipase per liter in the supernatant, 40 times more than the single-copy strain with the LIP2 promoter. This work provides a new expression system in I: lipolytica that results in strains devoid of bacterial DNA and in strains producing a high level of lipase for industrial uses, waste treatment, and pancreatic insufficiency therapy.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 27/11/20 alle ore 22:19:53