Leukotoxin gene of pasteurella haemolytica

Details Leukotoxin gene of pasteurella haemolytica Leukotoxin gene of pasteurella haemolytica

1986-11-26

1991-10-08

Weimar, Elizabeth C.

Chemistry: molecular biology and microbiology

Micro-organism, tissue cell culture or enzyme using process...

Recombinant dna technique included in method of making a...

435 91, 4351723, 4353201, 43525233, 4352351, 536 27, C12P 2102, C12P 1934, C12N 1500, C12N 121, C12N 700, C12N 1512

Patent

active

050554000

ABSTRACT:
The gene coding for Pasteurella haemolytica leukotoxin can be cloned in a plasmic expressed in Escherichia coli. The leukotoxin gene can be isolated from a clone bank of P. haemolytica. The clone bank is constructed by partial digestion of genomic DNA. The resultant 5 to 10 kilobase-pair fragments are ligated into plasmid vector pBR322. The resultant clones are screened for the production of P. haemolytica soluble antigens by a colony enzyme-linked immunosorbent assay blot method with a rabbit antiserum raised against the soluble antigens. The clones producing P. haemolytica soluble antigens are then analyzed for the production of the leukotoxin by a cytotoxicity assay with cells from a bovine leukemia-derived B-lymphocyte cell line as the target cells. Positive clones are identified, and subsequent restriction analysis of the recombinant plasmids shows the same insert DNA is cloned in the plasmid vector. The DNA sequence analysis of the insert DNA reveals regions coding for the leukotoxin.

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