›› 2011, Vol. 29 ›› Issue (5): 1-321-326.

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Construction of GCV-Specific Hammerhead Ribozyme Recombinant Vector of Alpha-8 Giardin in Giardia lamblia

 WEI  Chao-Jun, LU  Si-Qi, CAO  Li-Jing, TIAN  Xi-Feng   

  1. 1 Clinical Laboratory,Gansu Provincial People Hospital,Lanzhou 730000,China; 2 Department of Pathogen Biology,Capital Medical University,Beijing 100069,China;3 Pediatric Intensive Care Unit,Hebei Children′s Hospital,Shijiazhuang 050031,China; 4 College of Life Science,Hebei United University,Tangshan 063000,China
  • Online:2011-10-30 Published:2012-09-27

Abstract: Objective   To construct a GCV-ribozyme recombinant vectors of α-8 giardin in Giardia lamblia.  Methods   The secondary structure of α-8 giardin mRNA (GenBank Accession No. AY781323) was analyzed with the RNA draw software. According to the proportion of G ∶ C and principles of designing hammerhead ribozyme,suitable ribozyme cleavage points were chosen. A specific antisense-hammerhead ribozyme(H8)was designed and synthesized. The ribozyme was cloned into Giardia canis virus (GCV) vector to construct a recombinant viral vector-pGCV634/H8/1423. The vector was linearized and transcripted into the trophozoites of G. lamblia by electroporation method. The α-8 giardin mRNA level of the transfectants and normal trophozoites were analyzed 24 h after electroporation by RT-PCR.   Results   The recombinant vector of GCV-specific hammerhead ribozyme of α-8 giardin in Giardia lamblia(pGCV634/H8/1423) was constructed. RT-PCR assays showed the ribozyme (H8) mRNA can be detected 24 h after transfection and α-8 giardin mRNA was cleaved effectively by ribozyme (H8) introcellularly.   Conclusion   pGCV634/H8/1423 can transfect Giardia trophozoites and cleave mRNA of α-8 giardin intracellularly.

Key words: Giardia lamblia;Giardin;&alpha, -8 giardin;Hammerhead ribozyme;Giardia canis virus (GCV)