BIOLISTIC TRANSFORMATION OF A PROKARYOTE, BACILLUS-MEGATERIUM

被引:54
作者
SHARK, KB
SMITH, FD
HARPENDING, PR
RASMUSSEN, JL
SANFORD, JC
机构
[1] CORNELL UNIV,DEPT HORT SCI,GENEVA,NY 14456
[2] PLATTSBURGH STATE UNIV COLL,DEPT BIOL SCI,PLATTSBURGH,NY 12901
[3] UNIV NEBRASKA,SCH BIOL SCI,LINCOLN,NE 68588
关键词
D O I
10.1128/AEM.57.2.480-485.1991
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
We present a simple and rapid method for introducing exogenous DNA into a bacterium, Bacillus megaterium, utilizing the recently developed biolistic process. A suspension of B. megaterium was spread onto the surface of nonselective medium. Plasmid pUB110 DNA, which contains a gene that confers kanamycin resistance, was precipitated onto tungsten particles. Using a biolistic propulsion system, the coated particles were accelerated at high velocities into the B. megaterium recipient cells. Selection was done by use of an agar overlay containing 50-mu-g of kanamycin per ml. Antibiotic-resistant transformants were recovered from the medium interface after 72 h of incubation, and the recipient strain was shown to contain the delivered plasmid by agarose gel electrophoresis of isolated plasmid DNA. All strains of B. megaterium tested were successfully transformed by this method, although transformation efficiency varied among strains. Physical variables of the biolistic process and biological variables associated with the target cells were optimized, yielding > 10(4) transformants per treated plate. This is the first report of the biolistic transformation of a procaryote.
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页码:480 / 485
页数:6
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