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Background  Most enzymatic mutation detection methods are based on the cleavage of heteroduplex DNA by a mismatch-specific endonuclease at mismatch sites and the analysis of the digestion product on a DNA sequencer. Important limitations of these methods are the availability of a mismatch-specific endonuclease, their sensitivity in detecting one allele in pool of DNA, the cost of the analysis and the ease by which the technique could be implemented in a standard molecular biology laboratory.    Results  The co-agroinfiltration of ENDO1 and p19 constructs into  N. benthamiana  leaves allowed high level of transient expression of a mismatch-specific and sensitive endonuclease, ENDO1 from  Arabidopsis thaliana . We demonstrate the broad range of uses of the produced enzyme in detection of mutations. In human, we report the diagnosis of the G1691A mutation in  Leiden factor-V  gene associated with venous thrombosis and the fingerprinting of HIV-1 quasispecies in patients subjected to antiretroviral treatments. In plants, we report the use of ENDO1 system for detection of mutant alleles of  Retinoblastoma - related  gene by TILLING in  Pisum sativum  and discovery of natural sequence variations by Eco-TILLING in  Arabidopsis thaliana .    Conclusion  We introduce a cost-effective tool based on a simplified purification protocol of a mismatch-specific and sensitive endonuclease, ENDO1. Especially, we report the successful applications of ENDO1 in mutation diagnostics in humans, fingerprinting of complex population of viruses, and in TILLING and Eco-TILLING in plants.

Mutation detection using ENDO1: Application to disease diagnostics in humans and TILLING and Eco-TILLING in plants