MONOCLONAL-ANTIBODIES SPECIFIC FOR ELONGATION FACTOR-TU AND COMPLETE NUCLEOTIDE-SEQUENCE OF THE TUF GENE IN MYCOBACTERIUM-TUBERCULOSIS

被引:16
作者
CARLIN, NIA
LOFDAHL, S
MAGNUSSON, M
机构
[1] NATL BACTERIOL LAB,DEPT BACTERIOL,S-10521 STOCKHOLM,SWEDEN
[2] STATENS SERUMINST,TUBERCULIN LAB,DK-2300 COPENHAGEN,DENMARK
关键词
D O I
10.1128/IAI.60.8.3136-3142.1992
中图分类号
R392 [医学免疫学]; Q939.91 [免疫学];
学科分类号
100102 ;
摘要
Monoclonal antibodies against mycobacterial antigens were produced by immunizing LOU/C rats with live Mycobacterium bovis BCG. The antibodies were characterized by an enzyme-linked immunosorbent assay and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by Western blotting (immunoblotting). One antibody, MAMB 2, reactive with a 47-kDa protein was used to screen a lambda-gt11 M. tuberculosis gene library (R. A. Young, B. R. Bloom, C. M. Grosskinsky, J. Ivanji, D. Thomas, and R. W. Davis, Proc. Natl. Acad. Sci. USA 82:2583-2587, 1985). Three recombinant phages reactive with MAMB 2 in plaque lysates were isolated, and part of the insert was sequenced. The mycobacterial inserts were all expressed as proteins fused with beta-galactosidase when the phages were induced as lysogens in Escherichia coli. The entire M. tuberculosis tuf gene was obtained by screening the lambda-gt11 library with a DNA probe specific for the primary clones. A phage isolated from this screening was able to express the native protein in E. coli when introduced as a lysogen. A comparison of the entire gene sequence and the deduced protein sequence with the EMBL DNA and Swiss-Prot protein data libraries revealed strong homologies with elongation factors of bacteria, yeast mitochondria, and a plant chloroplast.
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页码:3136 / 3142
页数:7
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