Conditional gene knock-down by CRE-dependent short interfering RNAs

Short interfering RNAs (siRNAs) are short (21-23 nt) double-stranded RNAs that direct the sequence-specific degradation of corresponding mRNAs, resulting in suppression of gene activity. siRNAs are powerful tools for gene functional analysis in mammals. Chemically synthesized siRNAs permit transient gene repression but preclude inhibition of stable gene products as well as long-term phenotypic analyses. Permanent gene suppression can be achieved by transcribing siRNAs as stem-loop precursors from Pol III promoters. This approach, however, has a major limitation: inhibition cannot be controlled in a time- or tissue-specific manner. Thus, the approach cannot be applied to genes essential for cell survival or cell proliferation. To overcome these limitations, we have designed a CRE-lox-based strategy that allows one to repress gene activity in a time- dependent manner in cells, and in a time- or tissue-dependent manner in animals. Our approach promises to improve dramatically the procedures for functional genetics in mammals.