About the Knockout Mouse Project knockout strategiesThe two groups funded to produce knockout alleles for the KOMP project use targeting constructs that have different and complementary properties. Below is a general outline of these constructs that may be helpful in your assessment of the preferred approach for targeting your gene(s) of interest. Constructs used by Regeneron, Inc.Regeneron, Inc. will use the following general construct for creating knockouts for KOMP. In most cases these will be complete null alleles that delete the entire protein coding sequence of the target gene. This allele design can be applied to any gene transcribed by RNA polymerase II regardless of its size, intron-exon structure, RNA splicing pattern, or protein-coding capacity.  Reference: Valenzuela, D. M., Murphy, A. J., Frendewey, D., Gale, N. W., Economides, A. N., Auerbach, W., Poueymirou, W. T., Adams, N. C., Rojas, J., Yasenchak, J., et al. (2003). High-throughput engineering of the mouse genome coupled with high-resolution expression analysis. Nat Biotechnol 21, 652-659. Constructs used by the CSD (CHORI/Sanger/UC Davis) teamCSD: CHORI (Children's Hospital Oakland Research Institute), the Wellcome Trust Sanger Institute, and the University of California at Davis are using two constructs, both of which rely on identifying a key exon relevant to all transcripts of the gene. Each of these constructs can produce reporter knockouts, conditional knockouts, and null alleles. Thus, this construct is extremely flexible in the types of alleles that can be produced. However, the genes must be of sufficient size to contain a "differential" exon of note, and the approach might not be readily amenable for small genes with few exons. Promoterless Targeting Cassette:  Promoter Driven Targeting Cassette:  More detailed description of alleles that can be produced:  |
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