Historically, the rat has been used for mapping a variety of complex diseases and a multitude of genetic and genomic reagents exist to support these studies. RGD contains much of the raw data generated by various groups and individual researchers and this section describes how one might bring these to bear on a mapping study.

• How do I find what genes are causing a disease or phenotype?
• How do I find out if my gene has been mapped?
• How do I do a genetic mapping study?
• How do I select a strain for a mapping cross?
• How do I find polymorphic markers in my mapping cross?
• How do I find the software to make the maps and find the QTLs?
• How do I map my gene without mapping a cross?

• SSLPs, QTLS, Maps,
  References, Genes
• Genome Scanner
• RH Mapserver
• VCMap

For more background information:

1. Genetic Analysis of Inherited Hypertension in the Rat, John Rapp, Physiological Reviews, 90:135-172,2000
   Review article on mapping hypertension in the rat, also great introduction to mapping in general
2. Genome Analysis, A laboratory manual, Vol 4: Mapping Genomes. Birren, etal. CSHL Press.
   Excellent lab manual describing a wide variety of mapping techniques in humans and model organisms.
3. Analysis of Human Genetic Linkage, Ott, J. 3rd Edition 1999, JHU Press.
   'Human-centric' perspective but covers many fundamental techniques particularly statistical aspects.
4. Modern Genetic Analysis, Griffiths, etal [online at NCBI]
   Online textbook describing many fundamental analysis techniques

This is one of the fundamental questions being addressed by Rat research and RGD is designed to support many aspects of this work. Reference #1 (Rapp, Physiological Reviews) describes the process of determining the genes involved in hypertension and is a good overview of the techniques that can be applied to any disease or phenotypic trait.

Are any genes or genomic regions already related to my disease/phenotype?

RGD collects published information that relates genes or regions to particular diseases or phenotypes, this can be accessed in several ways:

1. Advanced Search- The functionality to search RGD using multiple text fields, to use operators (AND, NOT, OR) and to allow users to limit their search results to specific data types like genes, strains, QTLs, etc. Many data types have associated disease information included in the data reports. For example:
1. Genes have disease notes entered by the curation staff that contain published associations between a gene and phenotype
2. Quantitiative Trait Loci ( QTLs) define regions of the genome already believed to contain gene(s) associated with a particular disease or phenotype. The QTL search page lists all the phenotypes for which regions have been identified.
2. Disease Portals- RGD is developing disease portals to direct users to disease information of most interest to them.
3. Comparative mapping - If a gene is known to be associated with the disease/phenotype in another organism (e.g. Human or Mouse), it is possible that it plays a similar role in the Rat. RGD provides the VCMap tool, which facilitates moving between the rat, mouse and human genomes. This is the premise of comparative genomics and other databases such as MGI (Mouse) and Entrez Gene (Rat, Mouse, Human and more) provide gene/disease notes and QTL information on these other organisms that can be used to work back to Rat

RGD holds data from three main maps, two genetic maps (FHH x ACI v7, SHRSP x BN v7) and one radiation hybrid map (MCW/WI v2.1) and many genes, ESTs and SSLP (microsatellite) markers are mapped on these existing maps. These maps can be viewed in the Maps section of the database. RGD also stores cytogenetic map data where genes have been mapped by FISH or similar techniques. By searching for your gene or marker within RGD using the Advanced Search you can find out if it has already been mapped on one of these standard maps.

To find the Physical location of your gene/sequence in the Genome, please see How can I find the genomic (physical) location of....?.

You can also try Comparative Mapping if your gene has already been mapped in human and/or mouse. Approaches to this are covered under the Genomics section.

 Whilst a complete description is out of the scope of this document, there are certain steps that can be outlined here. The assumption is made that you are attempting to map the genetic component(s) of a phenotypic trait of interest.

• Your trait must have a measurable genetic component.
• You should identify a strain that exhibits the phenotypic trait (affected strain).
• Select a control strain that does not exhibit the trait (unaffected strain).
• Depending on the mode of inheritance of the trait, an appropriate cross is set up between the affected and unaffected strain (For more information see Ref #2, Chapter 2).
• The offspring are phenotyped for the trait(s) of interest and genotyped (typically using microsatellite markers) to determine the genetic makeup of the offspring - what sections of each parental genome are present in each offspring.
• Statistical techniques are used to correlate phenotype with genotype (see the software section below), ideally resulting in a significant correlation in a specific region, which is termed a quantitative trait locus.

For more description see "locating the genes" in Chapter 18 of Modern Genetic Analysis, also the Nature Reviews Genetics article "Mapping and analysis of QTL in experimental populations" by Rebecca Doerge (Nature Reviews Genetics 3, 43-52 (2002))

The easiest way is to search the RGD strain records for strains that exhibit your trait of interest. The strain records are searchable by keyword (example: Search strains for 'Hypertension') quickly indicating if a strain has been studied for that trait. Another method is to search the literature (at RGD or PubMed) for other researchers who have studied the same trait or disease in the rat and investigate which strains they used. If a QTL has been mapped for your trait, the RGD QTL report will list which strains were used in the cross.

To do this manually, one would run PCR reactions for a marker to be tested (usually microsatellite markers) looking for a difference in the product sizes between DNA from the affected and unaffected strains. If such a polymorphism is detected then that marker can be used in a genetic mapping cross. The polymorphism is essential to be able to determine which parental strain contributed to the genome of the sibs.

RGD has a tool designed to aid in this process, Genome Scanner. Based upon the microsatellite data derived from the ACP project (4500 markers measured in 48 common strains) it will list the polymorphic markers between two selected strains.

What if my strain of interest isn't part of the 48 strains? You may have to resort to the manual methods described above, alternatively, seek out other researchers who have used these strains and they may have a list of polymorphic markers for your cross. The references attached to the RGD Strain records provide a good resource, particularly if the paper is a QTL mapping paper.

Popular, free, software packages used in the creation of genetic maps and QTL mapping include:

• MapMaker3
• QTL Cartographer
• MapMaker/QTL
• MapManager suite: QT, QTX

More extensive list of software available at Rockefeller University.

Radiation Hybrid mapping can be used to efficiently map any segment of DNA for which a pair of unique PCR primers can be designed. This avoids the need for animal crosses when one simply wishes to find out where a gene/est/marker resides in the genome. The T55 radiation hybrid panel for the rat was sold by ResGen, however the future of this panel is currently unknown since ResGen has closed down. They currently have more information available, though actually purchasing the panel may or may not still be possible.

RH mapping follows these fundamental steps:

• Unique PCR primers are designed around the sequence of interest
• The primers are run against all clones in the RH mapping panel (there are 106 clones in the rat RH though typically only 94 are used, allowing the reactions to be done in a 96 well plate with rat and hamster controls)
• The reaction results are scored for each clone, 1 for the presence of product of the correct size, 0 for no product. 2 if the the results are uncertain. These scores are listed together as a vector, as shown below:
  aldoa 0000000210000000000000001000000000010020000010000000001010100110001012100010000021011101000100222222
• This vector is compared against an pre-existing framework map using software such as RHMapper. If the new vector can be placed against the framework with sufficient statistical support, the map location of the DNA segment will be determined.

The RGD RH Map Server can be used to place your RH vectors against the published MCW/WI RH framework map. This convenient solution relieves the researcher from having to create their own framework map and from having to install and learn complicated mapping software. However, sound understandings of the limitations of this technique are also required. Please refer to “Background Information” references numbers 2 and 4, chapter 6 in both, for a comprehensive introduction to RH mapping.