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MERLOT II


    

Learning Exercise


Material: PedigreeLab
Submitted by: Jeff Bell on Sep 04, 2000
Date Last Modified: Sep 04, 2000
Title: Mapping Human Genetic Traits
Description: Students map human disease genes using pedigrees and RFLPs.
Type of Task: Core activity, Student-centered, Team, Unsupervised
Time Required 3 hours
Topics: Linkage, mapping, human genetic diseases, RFLPs
Course: Intro Biology, Genetics
Audience: College Lower Div Major
Categories:
Learning Objectives: Demonstrate how pedigree analysis can be used to determine the mode of inheritance for a genetic disorder across several generations of humans.
Demonstrate how recombination data from genetic crosses can be interpreted and applied to discover the location of a gene on a chromosome and to develop chromosome maps.
Simulate the use of restriction fragment length polymorphism (RFLP) analysis to develop genetic maps of a chromosome.
Text of Learning Exercise:

Mapping Human Genetic Traits


Introduction


In FlyLab you studied modes of inheritance and chromosome maps in Drosophila melanogaster. In this laboratory you will have the opportunity to use pedigree analysis to simulate the inheritance of genes for human genetic disorders, and use RFLP analysis to study recombination in humans. Combining RFLP analysis with pedigree analysis will enable you to learn about, and understand, how the location of a gene can be assigned to a chromosome and you will pinpoint the location of a gene on a chromosome by using recombination data to generate a genetic map of that chromosome.


Objectives


The purpose of this laboratory is to:


  • Demonstrate how pedigree analysis can be used to determine the mode of inheritance for a genetic disorder across several generations of humans.
  • Demonstrate how recombination data from genetic crosses can be interpreted and applied to discover the location of a gene on a chromosome and to develop chromosome maps.
  • Simulate the use of restriction fragment length polymorphism (RFLP) analysis to develop genetic maps of a chromosome.

Assignments


Twenty-two different mutations of human genes that produce actual genetic diseases are included in PedigreeLab. Some of these mutations produce diseases that you have probably heard about and already studied, in other cases more rare genetic mutations and diseases are presented. You will use PedigreeLab to study a number of different pedigrees to develop a hypothesis to explain the mode of inheritance for the trait that you selected. Once you have done this, your goal is to study recombination between the mutant gene and markers that you will select, using RFLP analysis to help you determine the chromosome containing the mutant gene, and the location of the mutated gene relative to genetic markers on that chromosome. These assignments will be carried outusing the PedigreeLab.


Determining Inheritance Patterns Using Pedigrees


The first screen that appears in PedigreeLab is the Information view beginning with Alzheimer's Disease. Note that for each mutation you are provided with a short narrative of background information describing the history, phenotypes, and other symptoms created by the mutation. For this first excercise you just need to determine wether the inheritance of a particular genetic disease is autosomal dominant, autosomal recessive, X-linked dominant or X-linked recessive. Using pedigrees is much more difficult than crossing fruit flies becuase there are only a few children in each generation and you can't design your own crosses. Human geneticists use pedigrees from several different families to try to determine the inheritance pattern.



1. Determine the mode of inheritance of Familial type 3 Alzheimer Disease (AD3) (5 pts.)


Familial Alzheimer's is the default genetic disease but if you have changed the muatation go back to Alzheimer's by selecting gene AD3 in the pull down menu in the upper left corner (below the Information tab). Click on the "Full Pedigree" tab to view pedigrees from families with Familial Alzheimer's. The pedigree that is displayed on this page if one of 100 different sample pedigrees for AD3 available in PedigreeLab. To choose a different pedigree, click on one of up or down arrows in the upper right corner of the screen.
When studying these pedigrees, remember that circles represent females and squares represent males. Parents are connected by a red horizontal line. Black vertical lines from the parents indicate the offspring produced by these parents. Sibs are connected to each other by a horizontal black-lined bracket shown above the symbols. The symbol for an individual that expresses the mutantphenotypes is shaded blue. First generation parents are shown at the top of the
pedigree followed by the second generation, third generation etc. Look at several different pedigrees until you are confident that you have figured out the inheritance pattern for this disease. What is your hypothesis for the inheritance pattern? What is the evidence from the pedigrees that supports your hypothesis? Choose a simple pedigree that provides evidence for your hypothesis and assign genotypes to the individuals in the pedigree.


To assign a genotype to each individual, double click on the circle or the square for that individual. A new box should open prompting you to type in the genotype. Type in "Aa" or "aa" or "XAXa" (for X-linked traits) or XaY, etc., then click OK to close the genotype box. Double click on the square for the next individual and type in the appropriate genotype, then click OK to close the genotype box. The genotype for each individual should appear below each symbol. The labeled pedigree can be exported as a GIF file by clicking on the Export Pedigree button at the lower right corner of the screen. A new browser window with your labeled pedigree will appear. You can now print your pedigree from this window.


To make this a little easier for you all of the pedigrees in the pedigreeLab have complete penetrance. Looking at the description of Alzheimer's, why would you expect this disease not to have 100% penetrance?




2. Determine the mode of inheritance of Menkes Disease (ATP7A) (5 pts.)


Analyse several pedigrees for Menkes disease and determine the mode of inheritance as you did for Alzheimer's. Be sure to include a labelled pedigree and an explanation of the evidence that supports your hypothesis for the mode of inheritance.


Mapping the Locus for a Mutant Gene by Studying Large Family Pedigrees, RFLP Analysis, and Recombination


Identifying the chromosome that contains the mutation that you are studying and mapping the locus for this mutation using PedigreeLab involves your ability to apply what you learned about the mode of inheritance for the mutation, and your ability interpret the results of RFLP and recombination data. This process is part of the challenge and fun of PedigreeLab.


Using the Large Family function is the key to the gene mapping functions of PedigreeLab. This function simulates databases that present family pedigrees for individuals with different genetic disorders that have been subjected to RFLP analysis using one of 17 different probes for markers on human chromosomes. The pedigrees are arranged according to genotypes for the RFLPs; phenotypes are also shown with these pedigrees. The probes in PedigreeLab do not correspond to actual marker loci in human chromosomes.


You will then pick an appropriate pedigree that will allow you to look for recombination between the marker sequence of DNA recognized by the probe and the mutation. Based on recombination frequencies that you generate, and LOD scores, you will then decide whether the probe and mutation are linked to the same chromosome. If they are linked, you will then use the Chromosome View function to pinpoint the locus for your trait based on recombination frequencies. All of the traits presented in PedigreeLab will map to their actual chromosomes as found in real life. No shortcuts or hints are given to determine linkage groups, hence you must be a gene hunter using trial and error and analysis of your data to eliminate loci for unlinked probes and find linked probes that will help you pinpoint the locus for the mutant gene!


3. Determine the chromosomal location of the AD3 gene (10 pts.)






Notice that on the far left of this box you have four
popup menu boxes that you can use to select the trait and
marker combination for each homologue of a pair in a female
parent. Click on one of the upper two boxes, these allow you
to choose the genotype for the trait. Note that the dominant
allele for AD3 is shown in capital letters and the recessive
allele for AD3 is shown in lower case letters. Click on one
of the lower two boxes, these allow you to choose the
genotype for the probe (marker). Note that the genotype for
a homologue with a restriction site is shown as "pbq4+", and
the genotype for a homologue lacking a restriction site is
shown as "pbq4-."



It is important that you determine the correct linkage
combination for the parent who is a double heterozygote
(A/a, +/-) for the trait and restriction site. Do this for
the female parent. For the first homologue select a linkage
cshown as "pbq4-."



It is important that you determine the correct linkage
combination for the parent who is a double heterozygote
(A/a, +/-) for the trait and restriction site. Do this for
the female parent. For the first homologue select a linkage
c box you have four
popup menu boxes that you can use to select the trait and
marker combination for each homologue of a pair in a female
parent. Click on one of the upper two boxes, these allow you
to choose the genotype for the trait. Note that the dominant
allele for AD3 is shown in capital letters and the recessive
allele for AD3 is shown in lower case letters. Click on one
of the lower two boxes, these allow you to choose the
genotype for the probe (marker). Note that the genotype for
a homologue with a restriction site is shown as "pbq4+", and
the genotype for a homologue lacking a restriction site is
shown as "pbq4-."



It is important that you determine the correct linkage
combination for the parent who is a double heterozygote
(A/a, +/-) for the trait and restriction site. Do this for
the female parent. For the first homologue select a linkage
c unlinked on different chromosomes.



To use this function, click on the Genetic Calculator tab
at the top of the screen, then click on the popup arrow next
to the box labeled "Trait & Probe Autosomal, Linked" at
the top of this window. This popup menu allows you to design
genotypes for different individuals depending on whether you
think that the trait you are trying to map is an autosomal
or sex-linked trait, and whether or not this trait is linked
or unlinked to the probe for a given marker. Because you
knowfrom studying the full pedigrees that AD3 is inherited
as an autosomal dominant condition, select the "Trait &
Probe Autosomal, Linked" to design genotypes that would show
the AD3 trait and pbq4 marker as linked on an autosome.



Notice that on the far left of this box you have four
popup menu boxes that you can use to select the trait and
marker combination for each homologue of a pair in a female
parent. Click on one of the upper two boxes, these allow you
to choose the genotype for the trait. Note that the dominant
allele for AD3 is shown in capital letters and the recessive
allele for AD3 is shown in lower case letters. Click on one
of the lower two boxes, these allow you to choose the
genotype for the probe (marker). Note that the genotype for
a homologue with a restriction site is shown as "pbq4+", and
the genotype for a homologue lacking a restriction site is
shown as "pbq4-."



It is important that you determine the correct linkage
combination for the parent who is a double heterozygote
(A/a, +/-) for the trait and restriction site. Do this for
the female parent. For the first homologue select a linkage
cunlinked on different chromosomes.



To use this function, click on the Genetic Calculator tab
at the top of the screen, then click on the popup arrow next
to the box labeled "Trait & Probe Autosomal, Linked" at
the top of this window. This popup menu allows you to design
genotypes for different individuals depending on whether you
think that the trait you are trying to map is an autosomal
or sex-linked trait, and whether or not this trait is linked
or unlinked to the probe for a given marker. Because you
know from studying the full pedigrees that AD3 is inherited
as an autosomal dominant condition, select the "Trait &
Probe Autosomal, Linked" to design genotypes that would show
the AD3 trait and pbq4 marker as linked on an autosome.



Notice that on the far left of this box you have four
popup menu boxes that you can use to select the trait and
marker combination for each homologue of a pair in a female
parent. Click on one of the upper two boxes, these allow you
to choose the genotype for the trait. Note that the dominant
allele for AD3 is shown in capital letters and the recessive
allele for AD3 is shown in lower case letters. Click on one
of the lower two boxes, these allow you to choose the
genotype for the probe (marker). Note that the genotype for
a homologue with a restriction site is shown as "pbq4+", and
the genotype for a homologue lacking a restriction site is
shown as "pbq4-."



It is important that you determine the correct linkage
combination for the parent who is a double heterozygote
(A/a, +/-) for the trait and restriction site. Do this for
the female parent. For the first homologue select a linkage
c

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