Producing human lactoferrin by high-density fermentation recombinant Pichia pastoris.
- Author:
Ge YING
1
;
Shu-hua WU
;
Jing WANG
;
Xiao-dong ZHAO
;
Jian-ming CHEN
;
Xiao-guang ZHANG
;
Yun-de HOU
Author Information
- Publication Type:Journal Article
- MeSH: Cloning, Molecular; Fermentation; Humans; Lactoferrin; biosynthesis; genetics; Pichia; genetics; metabolism; Recombinant Proteins; biosynthesis
- From: Chinese Journal of Experimental and Clinical Virology 2004;18(2):181-185
- CountryChina
- Language:Chinese
-
Abstract:
BACKGROUNDTo evaluate expression of human lactoferrin gene by high-density fermentation in recombinant Pichia pastoris on the premise of maintaining its biological activities.
METHODSThe neutrophil was isolated from human peripheral blood and its total RNA was prepared. Full-length cDNA of human lactoferrin gene was then obtained by RT-PCR, cloned into expression vector pPIC 3.5 K and transformed into Pichia pastoris strain KM71. With two-layer filter method, the transformants with high-productivity of human lactoferrin were screened out into fed-batch high-density fermentation. And later, the physical, chemical and biological activities of fermentation product were detected preliminarily.
RESULTSThe strain p3.5-k-7 with better productivity of human lactoferrin was screened out into fed-batch high-density fermentation. The fermentation lasted nearly for nine days, with A-600 of culture once above 260 and the highest productivity of human lactoferrin being 115 mg/L, 7.67 times the amount of that in shake flask cultivation.
CONCLUSIONThe authors successfully realized high-density fermentation expression of human lactoferrin gene in recombinant Pichia pastoris.