Construction and screening of phage display single chain antibody library against Bursaphelenchus xylophilus cellulase / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 900-905, 2005.
Article
in Chinese
| WPRIM
| ID: wpr-237053
ABSTRACT
A phage display single-chain variable fragment (scFv) library against Bursaphelenchus xylophilus cellulase (BXC) was constructed and used to screen the specific antibodies binding to BXC. The total RNA was extracted from fresh spleens of BALB/C mice immunized with BXC. Gene fragments encoding VH and VL were amplified by RT-PCR and assembled into a single chain by overlapping PCR with a linker DNA encoding the peptide (Gly4Ser)3. The recombinant fragments were cloned into the phagemids (pCANTABSE) and electroporated into E. coli TG1. The recombinant phagemids were rescued by reinfection of helper phage M13K07. The repertoire of the phage display antibody was about 5 x 10(4). The specific antibodies against BXC were obtained after five rounds of affinity selection. The positive phage clone was used to infect E. coli HB2151. SDS-PAGE and western blot analysis showed that the soluble scFv antibodies expressed bound specifically to BXC. The studies laid foundation for quarantine and pathological study of Bursaphelenchus xylophilu.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Parasitology
/
Plant Diseases
/
Recombinant Proteins
/
Immunoglobulin Variable Region
/
Helminth Proteins
/
Cellulase
/
Cloning, Molecular
/
Electroporation
/
Peptide Library
/
Pinus
Type of study:
Diagnostic study
/
Screening study
Limits:
Animals
Language:
Chinese
Journal:
Chinese Journal of Biotechnology
Year:
2005
Type:
Article
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