How you would generate the knockout construct

You have been asked to investigate a specific human disorder, Crystal disease. The disorder is caused by a recessive mutation in the human gene A1. The disorder results in impaired vision, cognitive defects and early death. A short lived mutation has been isolated in butterflies which also causes neural degeneration and has been shows to affect the butterfly homologue of A1. A mouse homologue gene has been identified but is not cloned ( no one has isolated the gene sequence) and no mutations in the gene are known to exist. You are not a MD therefore cannot conduct human research. Two proposals has been submitted to you for evaluation. A. The first recommends using butterlies to conduct a suppressor screen for the phenotype associated with the mutation of A1. B. The second recommends generating a mouse model for Crystal disease. Explain how both would be conducted and provide a rationale for choosing one or the other approach. For A describe how you would perform the mutangenesis, what criteria you would use to identify suppressors, how would you clone and sequence the suppressor gene. For B explain how you would obtain the mouse sequence, how you would generate the knockout construct, how you would make the knock out mouse and how you would evaluate the reliability of the mouse model.

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