Cloning, Sequencing and Subcloning of cDNA Coding for GroupⅠAllergen of Dermatophagoides farinae

›› 2004, Vol. 22 ›› Issue (3): 13-175.

• 实验报道 • Previous Articles     Next Articles

Cloning, Sequencing and Subcloning of cDNA Coding for GroupⅠAllergen of Dermatophagoides farinae

YANG Qing-gui,LI Chao-pin
   

  1. School of Medicine,Anhui University of Science & Technology,Huainan,232001,China
  • Received:1900-01-01 Revised:1900-01-01 Online:2004-06-30 Published:2004-06-30

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Abstract: 　Objective To clone,sequence and subclone the cDNA coding for group 1 allergen of Dermatophagoides farinae(Derf1). Methods The cDNA of Derf1 was amplified by RT-PCR and PCR. After purified, the gene fragment was cloned into a vector pMD-18T. The recombinant plasmid pMD-18T-Derf1 was transformed into E.coli JM109. Positive clones were screened and identified by PCR and digestion with restriction enzyme. The sequence of inserted Def1 gene fragment was also detected. Derf1 was then subcloned into the vector of pET-32a(+). Results The Derf1 gene fragment of Dermatophagoides farinae was specifically amplified from RNA by RT-PCR and PCR. The recombinant plasmid pMD-18T-Derf1 and pET-32a(+)-Derf1 was constructed and digested by Bam HⅠand SacⅠ, the size of gene fragment was 646 bp and in accordance with the expected one. Conclusion The pET-32a(+)-Def1 subcloning has been constructed successfully.

Key words: Dermatophagoides farinae, Allergen, cDNA, Sequence analysis