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Construction and identification of Fas-targeting siRNA-expressing plasmid.

ABSTRACT:

Objective

To study the therapeutic potential of Fas inhibition in different diseases, a Fas-targeting siRNA (small interfering)-expressing plasmid was constructed.

Methods

The U6 promoter cassette and siFas (small interfering RNA that inhibit Fas expression) template sequence were obtained by PCR method. They were cloned into modified pcDNA3.1. The resultant plasmid pU6-siFas was transfected into P815 cells with lipofectin2000 and selected under G-418-containing culture medium. Fas inhibition in stably transfected cells was detected by immunocytochemistry.

Results

The plasmid pU6-siFas efficiently reduced the expression of Fas and conferred G-418 resistance in P815 cells.

Conclusion

The successful construction of the siRNA expressing plasmid will facilitate the application of RNA interference technique and lay the foundation for further study of Fas inhibition in the treatment of different diseases such as aplastic anemia and acute liver failure.

SUBMITTER: Liu SH

PROVIDER: S-EPMC1389803 | biostudies-literature | 2005 Jul

REPOSITORIES: biostudies-literature

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Construction and identification of Fas-targeting siRNA-expressing plasmid.

Liu Su-hu SH Zhang Wang-gang WG Zhang Mei M Zhu Qing Q Tian Wei W

Journal of Zhejiang University. Science. B 20050701 7

<h4>Objective</h4>To study the therapeutic potential of Fas inhibition in different diseases, a Fas-targeting siRNA (small interfering)-expressing plasmid was constructed.<h4>Methods</h4>The U6 promoter cassette and siFas (small interfering RNA that inhibit Fas expression) template sequence were obtained by PCR method. They were cloned into modified pcDNA3.1. The resultant plasmid pU6-siFas was transfected into P815 cells with lipofectin2000 and selected under G-418-containing culture medium. Fa ...[more]

PMID: 15973771

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