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Publication Date
1997
Location
Manitoba and Saskatchewan
Description
A complementary DNA (cDNA) library was constructed from the mRNA of adult worms of Fasciola hepatica in the expression vector gt11, the size of the library contained approximately 2.4 x 105 recombinants and the recombinant efficiency was 85%. The library was directedly screened with a rabbit antisera raised against a excretory-secretory (ES) antigen of Fasciola hepatica. Two positive clones of strong signal were selected from a group of 200 positive clones, named FH3 and FH7. The length of external fragments of two recombinants were 0.85Kb (FH3) and 1.15Kb (FH7). The E.coli 1089 host was infected by recombinants FH3 and FH7 respectively, through SDS - PAGE (polyacrylamide gel electrophoresis) after induced by isopropyl-D-thiogalactopyranoside (IPTG) (final concentration 5mM), the results indicated that the molecular weight of fusion protein expressed (-galactosidase fusion protein) of recombinant FH3 was 138KD, and that of FH7 was 156KD, the rate of fusion proteins expression was approximately 100mg/liter. Western - blot showed that two hybrid proteins corresponded to 26.2KD protein band ((FH3) and 38.8KD protein band (FH7) of Fasciola hepatica ES antigens.
Citation
Heng, Xu; Li, Zheng; Shigui, Liu; Bin, Shen; and Fend, Wu, "Genomic Cloning of Fasciola Hepatica Excretory-Secretory Antigenic Genes in E. Coli" (1997). IGC Proceedings (1985-2023). 38.
(URL: https://uknowledge.uky.edu/igc/1997/session11/38)
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Genomic Cloning of Fasciola Hepatica Excretory-Secretory Antigenic Genes in E. Coli
Manitoba and Saskatchewan
A complementary DNA (cDNA) library was constructed from the mRNA of adult worms of Fasciola hepatica in the expression vector gt11, the size of the library contained approximately 2.4 x 105 recombinants and the recombinant efficiency was 85%. The library was directedly screened with a rabbit antisera raised against a excretory-secretory (ES) antigen of Fasciola hepatica. Two positive clones of strong signal were selected from a group of 200 positive clones, named FH3 and FH7. The length of external fragments of two recombinants were 0.85Kb (FH3) and 1.15Kb (FH7). The E.coli 1089 host was infected by recombinants FH3 and FH7 respectively, through SDS - PAGE (polyacrylamide gel electrophoresis) after induced by isopropyl-D-thiogalactopyranoside (IPTG) (final concentration 5mM), the results indicated that the molecular weight of fusion protein expressed (-galactosidase fusion protein) of recombinant FH3 was 138KD, and that of FH7 was 156KD, the rate of fusion proteins expression was approximately 100mg/liter. Western - blot showed that two hybrid proteins corresponded to 26.2KD protein band ((FH3) and 38.8KD protein band (FH7) of Fasciola hepatica ES antigens.
