Construction of Phage Display cDNA Library from Adult Worms of Schistosoma japonicum
- VernacularTitle:日本血吸虫成虫噬菌体展示cDNA文库的构建
- Author:
Yi SUN
;
Renchu JIA
;
Jinming LIU
;
Chunxiu YUAN
;
Yaojun SHI
;
Ke LU
;
Zhiqiang FU
;
Huan SUN
;
Youmin CAI
;
Jiaojiao LIN
- Publication Type:Journal Article
- Keywords:
Schistosoma japonicum;
Phage display cDNA library;
Adult worm
- From:
Chinese Journal of Parasitology and Parasitic Diseases
1997;0(05):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To screen protective antigen genes and construct the T7 phage display library from adult worms of Schistosoma japonicum.Methods Total RNA was extracted from adult worms of S.japonicum by Trizol reagent and mRNA was isolated from the total RNA.The ds cDNA was synthesized by reverse transcription using random primer.Directional EcoRⅠ/HindⅢ linkers were ligated into the ends of ds cDNA and the ds cDNA was digested with EcoRⅠand HindⅢ,which resulted in ds cDNA with EcoRⅠand HindⅢ adhering ends.The digested ds cDNA fragments longer than 300 bp in length were fractionated and ligated into T7 Select 10-3b vector.After packaging in vitro,the T7 Select 10-3b vector was transformed into BLT5403 to construct the T7 phage display cDNA library.Plaque assay and PCR were used to evaluate the library.Seven known objective genes of S.japonicum were screened by PCR to detect the representation of the library.Result Primary library capacity was 4.98?106 pfu,and the titer of amplified library was 3.85?1011 pfu/mL.The PCR identification result of 96 clones picked at random showed that recombination rate was 93.8%,in which 95.6% inserted cDNA fragments were longer than 300 bp in length.All the seven known objective genes of S.japonicum were amplified from the library.Conclusion The T7 phage display library from adult worms of Schistosoma japonicum was constructed.
