A general method for gene knockdown in mice by using lentiviral vectors expressing small interfering RNA.
We describe the use of lentiviral vectors expressing small interfering RNAs (siRNAs) to knock down the expression of specific genes in vitro and in vivo. A lentiviral vector capable of generating siRNA specific for GFP after transduction of 293T-GFP cell lines showed no GFP fluorescence. Furthermore, no GFP-specific RNA could be detected. When eggs from GFP-positive transgenic mice were transduced with lentivirus-expressing siGFP virus, reduced fluorescence could be seen in blastocysts. More interestingly, pups from F(1) progeny, which expressed siGFP, showed considerably diminished fluorescence and decreased GFP. We propose that an approach of combining transgenesis by lentiviral vectors expressing siRNAs can be used successfully to generate a large number of mice in which the expression of a specific gene(s) can be down-regulated substantially. We believe that this approach of generating "knockdown" mice will aid in functional genomics.
Tiscornia
Gustavo
G
Laboratory of Genetics, The Salk Institute, 10010 North Torrey Pines Road, La Jolla, CA 92037, USA.
Singer
Oded
O
Ikawa
Masahito
M
Verma
Inder M
IM
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D005260:Female; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D008164:Luminescent Proteins / Q000235:genetics; D008297:Male; D051379:Mice; D018345:Mice, Knockout / Q000235:genetics*; D016133:Polymerase Chain Reaction; D034741:RNA, Small Interfering / Q000235:genetics*
D017931:DNA Primers; D008164:Luminescent Proteins; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins