JAX® Gene Mapping Service details
Crossing appropriate strains
To optimize marker density, the strain you cross to should be as unrelated as possible to the mutant strain. We have pre-established marker panels for some of the more common strain combinations, such as C57BL/6J and 129S1/SvImJ, C57BL/6J and DBA/2J, and DBA/2J and BALB/cJ. We can develop new marker panels for other combinations.
Intercross or backcross?
Intercross progeny each represent two informative meioses. If you have plenty of mouse room space, you can use an intercross. If the mutation is recessive and does not affect reproduction or birth, you can produce about 80 F2 progeny, out of which 20 or so should be affected (homozygous recessive). For gene mapping, supply us with tail samples from 20 offspring, at least two of which are unaffected, and at least 10 of which are affected (depending on what you have available). The two unaffected serve as important controls. More affected animals maximize the resolution of the result.
For a dominant mutation, the unaffected animals are more informative. Please provide as close to 10 unaffected and 10 affected F2 from a dominant mutation as you have available.
Backcross progeny each represent a single informative meiosis. If you have a limited amount of mouse room space, you can use a backcross. Assuming the mutation is recessive and does not affect reproduction or birth, you can produce about 40 N2 progeny (F1 backcrossed to affected parent if mutation is recessive), out of which 20 or so should be affected (homozygous recessive). Supply us with tail samples from all 40 offspring for gene mapping to approximate the same resolution as from 20 F2.
For a dominant mutation, the N2 generation will be produced from F1 backcrossed to wild type parent. The same 40 offspring of both phenotypes can be used for the mapping.
If you supply us with fewer samples than we request, the mapping resolution may be reduced.
Number of markers in the first and second scans
The number of markers used determines map resolution. We use 100-120 SNP markers in the first (genome-wide) scan. We select only informative markers for analysis, maximizing information return for your dollar. This scan will localize the mutation to a sub-chromosomal region. The exact size of the resulting candidate interval will depend on the chance location of key crossover events among the samples provided. A scan of 100 markers has average marker spacing of 30 MB. Thus, approximately 30 MB is the minimum size of a candidate region defined by this approach.
This level of low resolution mapping may be adequate for determining if a mutation is likely to be a remutation at a previously known locus or is a mutation in a novel gene. The information from a first pass scan may also be useful to identify flanking markers for breeding and for future experiments to more finely map the mutation.
If desired, we can follow up with a smaller secondary mapping scan focused on the candidate chromosomal region. The resolution of a secondary scan will depend on the available markers, the number of samples available for testing, and the chance location of defining crossover events. The secondary mapping will narrow the candidate region substantially, but is unlikely to pinpoint a single candidate gene. The cost of a secondary scan will depend on the number of available informative markers and the number of samples, but is usually significantly less than the cost of the original genome-wide scan.
Currently, JAX does not offer services to fine map/positionally clone a gene. Positively identifying a gene requires breeding additional animals with crossovers in the candidate region and fine mapping these with locally informative markers, or sequencing the entire candidate region or coding parts of the candidate region via sequence capture and deep sequencing.
Tail samples or processed DNA?
We prefer that you send us tail samples. Send them on dry ice, individually labeled with informative id numbers for each mouse.
We will use DNA that you supply if it is compatible with our process. Highly purified and correctly quantitated DNA is more likely to work. A 2mm tail snip should yield enough DNA for analysis. The Qiagen extraction kit or another comparable extraction method is preferred. The ideal DNA concentration is ~100-200ng/ul. DNA should be provided in 10mMTrisHCl or ddH20. (Tris EDTA (TE) should NOT BE USED as a storage buffer. EDTA inhibits the SNP reaction and should not be present in a concentration greater than 0.1 mM.)
Shipping the samples
Shipping all samples at the same time reduces your shipping and our processing costs.
Appropriate mapping projects
If a phenotype is contributed by multiple unlinked loci, the method for mapping single gene traits will not work. Mapping may also fail if multiple similar or related phenotypes are segregating in the cross and being considered together. Cases of significantly reduced penetrance can also prevent discovery of the underlying gene location. If you need help in determining if any of these issues may be affecting your cross, please inquire before submitting a project.
Multi-integrated transgenes
Transgenes can integrate into multiple locations. Although we can identify a multi-integrated transgene, we cannot finely resolve its location. If our analysis indicates that the transgene has integrated into multiple locations, we will contact you immediately.
