Mutation/techniques illustrated
For Case A: The DNA sequence files are from the human MP6d9 gene, GenBank accession #M37523, 383 bp, which is linked to the cystic fibrosis gene on chromosome 7. This simulates a PCR-amplified region near the CF gene. The mutated sequence files were created by replacing the G to A base change at nucleotide 170 which causes the loss of an Msp I site. No Southern blotting is necessary to observe the mutation.
RFLP marker outside of the actual disease gene, but genetically linked to the disease gene.
Reference: Huth,A., et al., Nucleic Acids Res. 17 (17), 7118, 1989.
For Case B: The DNA sequences are from the human CFTR gene, GenBank accession #M28668, 6129 bp. The mutated sequence files were created by deleting nucleotides 1653-1655 (delTTC). The normal probe is 25 bp spanning this region, while the CF mutation probe contains the TTC deletion.
Reference: Riordan et al., Science 245:1066-1073, 1989.
Case A:
As Sharon Brown browsed the local newspaper, she noticed the story about the five-year old boy with cystic fibrosis who lives on the next block. The article was mainly a human interest story about how the family was coping. There also was some background information about the disease and its inheritance patterns, including the statistics indicating that approximately 1 in 18 people in this part of Minnesota carried a cystic fibrosis mutation.
Sharon is two months pregnant. She realizes that she and her husband, Bob, should have been tested for the cystic fibrosis (CF) mutation since they each have some family history of the disease, but they really hadn’t expected to have a child so soon. She discusses this with her physician during her check-up the next day, and together they decide to test Sharon and Bob for a mutation in linked to the CF gene. They also decide to test the developing fetus. Two other families in the same town who also are in the first trimester of a pregnancy, Jill and Mike Jones and Carol and Ron Smith, also decide to be tested after reading the article.
- Brown family:
- Bob – homozygous normal
- Sharon- heterozygous
- Fetus – heterozygous
- Jones family:
- Jill – heterozygous
- Mike – heterozygous
- fetus – homozygous CF mutation
- Smith family:
- Carol – heterozygous
- Ron – heterozygous
- Fetus – homozygous normal
- Controls on all three gels:
- DNA heterozygous control
- DNA homozygous mutation control
- DNA homozygous normal control
Case B: (Contributed by Stephanie Dahlby, Dan Tally, and Janelle Veerkamp, students at UW-River Falls)
Lynda and Jim are expecting their first child. Recently, however, they learn that Lynda’s aunt died of CF and Jim’s uncle died of CF. They are worried that they might be carriers for the disease and pass cystic fibrosis on to their unborn child. They learn about a procedure which can determine whether they are carriers. They also learn about a procedure called amniocentesis which can detect if their unborn child has CF or is a carrier. However, amniocentesis is a very risky procedure. Jim and Lynda ultimately decide that they first want to be tested to see if they are carriers for the disease. If they learn that they both are carriers, they would like to go through with the amniocentesis to see if their child is affected.
- Lynda – heterozygous
- Fetus – homozygous CF mutation
- Jim – heterozygous
- Control DNA with F508 mutation
- Control normal DNA, without mutation
Case C:
The pre-marriage counseling session Carl and Maggie are having with Pastor Frank is not going at all as they had expected it to. After some of the anticipated discussion of relationship issues, the conversation turns to family planning. When both Carl and Maggie say they want to have children, Pastor Frank, instead of giving advise on how to properly rear children, begins to talk about genetic testing for Cystic Fibrosis! It turns out that Pastor Frank and his wife had two children affected with CF who died in their early teens. Because of the relatively high frequency of CF carriers and his opposition to abortion, Pastor Frank believes that all couples should be tested for the CF gene before getting married. Carl and Maggie are not sure they share Pastor Frank’s beliefs but decide to go along with being tested.
- Carl – heterozygous
- Maggie – heterozygous
- Control DNA with F508 mutation
- Control normal DNA without mutation