Acta Academiae Medicinae Sinica

Acta Academiae Medicinae Sinica

Acta Academiae Medicinae Sinica ›› 2009, Vol. 31 ›› Issue (6): 760-764.doi: 10.3881/j.issn.1000-503X.2009.06.022

• Original Articles • Previous Articles     Next Articles

Construction and Identification of the Lentiviral Vector of Repressor Element-1/Neuron-restrictive Silencer Element Double-stranded RNA

LI Hong-tu1,2;LIU Xiao-yu1; PANG Xi-ning1   

  1. 1Department of Developmental Biology, Key Laboratory of Cell Biology of Ministry of Public Health,China Medical University, Shenyang 110001, China
    2Key Laboratory of Reproduction Health of Liaoning Province, Research Institute of Family Planning ofLiaoning Province, Shenyang 110031, China
  • Received:2009-09-08 Revised:1900-01-01 Online:2009-12-30 Published:2009-12-30

Abstract: ABSTRACT:Objective To construct a lentiviral vector of repressor element-1/neuron-restrictive silencer element (RE-1/NRSE) double-stranded RNA (dsRNA). Methods The RE-1/NRSE cDNA containing both sense and antisense oligo DNA fragments of the targeting sequence was synthesized and cloned into the pGC-LV vector. The obtained lentiviral vector containing RE-1/NRSE dsRNA was confirmed by PCR and sequencing. A total of 293T cells were cotransfected with lentiviral vector of L-smNRSE/RE-1, pHelper 1.0, and pHelper 2.0. The titer of virus was measured based on the expression level of green fluorescent protein. The transfection efficiency of green fluorescent protein into rat mesenchymal stem cells was calculated. Results PCR and DNA sequencing demonstrated that the constructed lentivirus vector of L-smNRSE/RE-1 produced RE-1/NRSE dsRNA.The titer of the concentrated virus was 4×108 TU/m1. The virus was stably transfected into rat mesenchymal stem cells, and the infection efficiency reached 100% when the multiplicity of infection was 80. ConclusionThe lentivirus vector of RE-1/NRSE dsRNA is successfully constructed.

Key words: repressor element-1/neuron-restrictive silencer element, double-stranded rna, lentivirus vector