Microbial Status and Genetic Evaluation of Mice and Rats Proceedings of the 1999 US/Japan Conference (2000) / Chapter Skim
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A Phenotype-driven Approach to the Molecular and Functional Analysis of the Mouse Genome
A Phenotype-driven Approach to the Molecular and Functional Analysis of the Mouse Genome
Pages 105-115
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From page 105... ...
is employing chemical mutagenesis and broad-based phenotype screening to recover mutations targeted to specific chromosome regions for ascertaining the whole organism functions of mouse genes. Our strategy of chemical mutagenesis results in pedigrees of mice, each harboring a different DNA mutation for one of the many genes contained in the chromosome region.
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From page 106... ...
The current program has benefited from our hands-on experience as we increase both the chromosomal region target sizes and our scope and capacity for examining mice for as many different kinds of abnormalities as possible. Our current experiments take advantage of deletion screens, as described above, and more powerful methods utilizing chromosomal inversions as tools to make newly mutagenized chromosomes homozygous all without molecular genotyping.
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From page 107... ...
We know that mutagenesis and phenotype screening within different regions will result in the recovery of more or fewer mutations due to gene density differences in the regions, and/or to the proportion of genes in the regions that can mutate to a visible or lethal phenotype. Thus, whereas gene density is fixed, we should be able to affect mutation recovery rate by expanding the number and kinds of phenotype screens to increase the proportion of genes for which we can detect a mutation.
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From page 108... ...
Because we do have multiple test class animals in every pedigree to screen, we can rely on replication of any abnormality; and with heritability testing, false positives are minimized, even with highly variable traits. We are also certain, by visual genotyping, that we are performing our screens only on test class mice that can be expected to exhibit recessive mutant phenotypes, thus maximizing throughput and minimizing cost.
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From page 109... ...
However, by employing multiple tests, we can develop a pattern of response across tasks to inform us about a particular process. For example, a mouse with heightened anxiety but no memory deficit may perform poorly in the Y-maze spontaneous alternation memory task but show improved performance in the conditioned freezing memory task.
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From page 110... ...
to automatically compute relative dwell time for each user-defined portion of the open-field chamber. This is followed by assessment of acoustic startle response, prepulse inhibition of startle (PPI)
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From page 111... ...
Sperm will also be cryopreserved in the event that later recovery of the pedigree through in vitro fertilization is required. Using fluid or tissue samples from the test class allows efficient use of test class mice inasmuch as samples from any mouse can feed a large number of tests; we also archive a variety of tissues from each pedigree for future primary or secondary screens.
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From page 112... ...
for primary and secondary screening programs that greatly increase our opportunities for realizing the highest possible mutation recovery rate from our mutagenesis program. Statistical Analysis and Flagging Mutant Mice We use the test results from all other test class animals from the same mutagenesis experiment as controls inasmuch as all mice are handled the same and the majority of pedigrees yield normal test class progeny for any given phenotype.
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From page 113... ...
all primary data are currently entered into Excel spreadsheets, and from there will become part of a laboratory information system under development by ORNL's Computational Biosciences/Bioinformatics group. SUMMARY The goal of this large-scale mouse mutagenesis and phenotype screening program is to annotate DNA sequence with experimentally derived functional information about how individual genes perform in the context of a whole mammalian organism.
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From page 114... ...
1997. Chromosomal deletion complexes in mice by radiation of embryonic stem cells.
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From page 115... ...
Making more animals in that third generation so that you can breed it to make a fourth generation of multiple test class animals will limit the number of overall pedigrees that you can screen, which you do not want to do because only a certain percentage of the pedigrees are mutants in a region, or are mutant to a phenotype.
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