Linkage analysis tests for co-segregation of a chromosomal region and a trait locus of interest. In parametric linkage analysis, a specific disease model is used to describe segregation of the trait locus. In this section, we will walk through a parametric linkage analysis using MERLIN.

For this example, we will use a simulated data set that you will find in the examples subdirectory of the MERLIN distribution or in the download page.

The dataset consists of a 10-cM scan of candidate chromosome in a single pedigree where a rare dominant disorder is segregating (the pedigree is picture above). Ten microsatellite markers, each with 4 equally frequent alleles, were genotyped in all pedigree members. The genotypes and phenotypes are described in 3 files, a data file (parametric.dat), a pedigree file (parametric.ped) and a map file (parametric.map). An overview of MERLIN input files is available elsewhere.

The recommended first step in any analysis is to verify that input files are being interpreted correctly. So let's start by running pedstats... Pedstats requires an input data file (-d parameter) and pedigree file (-p parameter):

prompt> pedstats -d parametric.dat -p parametric.ped

By examining the abbreviated pedstats output below, you should be able to confirm that there is a single pedigree, with a total of 16 individuals (8 of these individuals are affected), and that there is no missing phenotype or genotype data.

Pedigree Statistics - 0.5.4
(c) 1999-2005 Goncalo Abecasis, 2002-2005 Jan Wigginton

The following parameters are in effect:
                 Pedigree File :  parametric.ped (-pname)
                     Data File :  parametric.dat (-dname)

PEDIGREE STRUCTURE
==================

       Individuals: 16
          Founders: 5 founders, 11 nonfounders
            Gender: 6 females, 10 males
          Families: 1

       Generations
           Average: 3.00 (3 to 3)
      Distribution: 3 (100.0%), 0 (0.0%) and 1 (0.0%)

AFFECTION STATISTICS
=====================

                    [Diagnostics]      [Founders] Prevalence
VERY_RARE_DISEASE       16 100.0%        5 100.0%      50.0%
            Total       16 100.0%        5 100.0%

MARKER GENOTYPE STATISTICS
===========================

                    [Genotypes]      [Founders]     Hetero
           MRK1       16 100.0%        5 100.0%      87.5%
           MRK2       16 100.0%        5 100.0%      75.0%
        (...statistics for other markers would appear here...)
          MRK10       16 100.0%        5 100.0%      75.0%
          Total      160 100.0%       50 100.0%      78.1%

The pedigree and data file seem to be okay. In addition to the standard Merlin input files, parametric linkage analyses require disease locus parameters to be specified in a separate text file. This text file has one row for each of the disease models to be evaluated, and can include as many different models as available memory allows. For this analysis, the file parametric.model specifies a single rare dominant disease model. Here are its contents:

In general, the file should be tab or space delimited, with 4 fields: affection status label (matching the data file), disease allele frequency, probability of being affected for individuals with 0, 1 and 2 copies of the disease allele (penetrances), and finally a label for the analysis model. A header line is included in the table above, for readability, but is not required. This file can also specify penetrance functions that depend on a covariate, such as age.

Okay ... let's run merlin! We will need to specify an input data file (-d parameter), pedigree file (-p parameter) and map file (-m parameter) as well as the file with trait model parameters (--model command line option). Since parametric linkage LOD scores tend to dip at marker locations, we will request an analyses at three equally spaced locations between each consecutive pair of markers with the --step 3 option. With all these options, the command line will look like this:

prompt> merlin -d parametric.dat -p parametric.ped -m parametric.map --model parametric.model --step 3

After running the command, you should first see the MERLIN banner and a summary of currently selected options:

MERLIN DEMO-VERSION - (c) 2000-2005 Goncalo Abecasis

The following parameters are in effect:
                     Data File :  parametric.dat (-dname)
                 Pedigree File :  parametric.ped (-pname)
                      Map File :  parametric.map (-mname)
            Allele Frequencies : ALL INDIVIDUALS (-f[a|e|f|file])

Data Analysis Options
           General : --information, --likelihood, --model [parametric.model]
         Positions : --steps [3], --maxStep, --minStep, --grid, --start,
                     --stop

Notice that allele frequencies were estimated by counting among all individuals (the default). In this case, this does not matter because all founders are genotyped. In practice, when analysing small datasets such as this one, it might be a good idea to genotype additional unrelated individuals to obtain better estimates of allele frequencies or to use an allele frequency file with custom frequencies.

After a minute or two, you should see analysis results at each location:

Parametric Analysis, Model Dominant_Model
=======================================================
       POSITION        LOD      ALPHA       HLOD
            (... some results edited to save space ...)
         35.000     -1.291      0.000      0.000
         37.500      2.037      1.000      2.037
         40.000      2.263      1.000      2.263
         42.500      2.358      1.000      2.358
         45.000      2.388      1.000      2.388
         47.500      2.201      1.000      2.201
         50.000      1.959      1.000      1.959
         52.500      1.585      1.000      1.585
         55.000     -9.291      0.000      0.000
            (... results continue at other locations...)

Each row indicates the estimated multipoint LOD score at a particular location. This is followed by the estimate proportion of linked families (since there is only one informative family in this sample, the proportion will always be 0.000 or 1.000), and the corresponding maximum heterogeneity LOD score. In this case the maximum LOD score of 2.407 is observed at position 45.000, the position of marker MRK5 in the map file.

Useful options for parametric linkage analyses options include requesting output with marker names, instead of cM positions (--markerNames option), requesting analysis along a grid of equally spaced locations (--grid n for an n-cM grid) rather than at a fixed number of steps between markers (--steps n for n-steps between consecutive markers), or requesting a graph summarizing results (--pdf). Try them out! For example...

prompt> merlin -d parametric.dat -p parametric.ped -m parametric.map --model parametric.model --grid 1 --markerNames --pdf

... would calculate the parametric LOD scores for a 1-cM grid along the chromosome and generate a PDF file with the resulting statistics.

That is it! That is all you need to get started with parametric linkage analysis in Merlin. Remember to set your disease model carefully, as an appropriate and careful choice of disease model is essential for parametric linkage analyses.

To learn about other analyses options, you might want to check the non-parametric linkage analysis section to find out how to conduct affecteds only linkage analyses. Or you could proceed to the error detection (improves power!), haplotyping, simulation or ibd estimation sections.