In search of a cancer gene
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In search of a cancer gene. You will search for a gene by Pedigree Analysis Gel electrophoresis Sequencing Data. Pedigree of “Valerie’s” family (3-130) Label Photo with typed figure legend and data interpretation ID “children” that carry mutation from DNA sequences.

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Presentation Transcript

In search of a cancer gene2 l.jpg

You will search for a gene by

Pedigree Analysis

Gel electrophoresis

Sequencing Data

Pedigree of “Valerie’s” family (3-130)

Label Photo with typed figure legend and data interpretation

ID “children” that carry mutation from DNA sequences

In search of a cancer gene




What does the pedigree of a li fraumeni patient look like l.jpg
What does the pedigree of a Li-Fraumeni patient look like? are:

  • A sarcoma patient is included

  • At least two immediate relative with other cancers before the age 45

  • As well as multiple cancers in other family members

  • Your lab manual provides a typical LF pedigree


Which gene is involved in the li fraumeni syndrome l.jpg
Which gene is involved in the Li-Fraumeni syndrome? are:

  • The p53 gene

  • F.P. Li, J.F. Freemeni, Jr.: Soft-tissue sarcomas, breast cancer and other neoplasm's: A familial syndrome? Annals of Internal Medicine, Philadelphia, October 1969, 71 (4): 747-752.


What are some characteristics of p53 l.jpg
What are some characteristics of p53? are:

  • “p” arm of chromosome 17

  • nuclear phosphoprotein

  • transcription factor

  • tumor suppressor gene

  • “hot spots”

    • Exons 5-8 are hot spots


P53 mutations l.jpg
P53 Mutations are:

  • Point Mutations

  • Mutation alters digest pattern


Identifying li fraumeni in three ways l.jpg
Identifying Li-Fraumeni in three ways: are:

  • Family History: leads to a pedigree

  • Germline mutations: restriction digests, followed by gel electrophoresis

  • Hot Spots: sequencing



The first step will be assign a pedigree to family containing patient valerie l.jpg
The first step will be assign a pedigree to family containing patient “Valerie”

  • Use the appropriate pedigree symbols as

    Outlined on 3-120.

  • For example:

Female free of cancer

Male free of cancer


The second step is to identify patterns of the restriction digest for patient valerie l.jpg
The second step is to identify containing patient “Valerie”patterns of the restriction digest for patient Valerie.


Step 2 restriction digest analysis of patient valerie l.jpg
Step 2: Restriction Digest Analysis of Patient: “Valerie”

  • Ladder

  • Wild Type DNA (gold standard)

  • Patient Peripheral Blood WBC DNA

  • Patient Tumor DNA

  • Patient Non-Tumor Breast Tissue


Step 3 identify the hot spot in valerie s children l.jpg
Step 3: Identify the “hot spot” in “Valerie”Valerie’s children.

  • Was this mutation inherited by her children?

  • Read the sequences of the 5 films

    • Film 1 Justin age 16

    • Film 2 Sheila age 14

    • Film 3 Robert age 10

    • Film 4 Angela age 8

    • Film 5 Anthony age 6


Summary today you will l.jpg
Summary: Today you will… “Valerie”

  • perform gel electrophoresis

  • Read sequencing films

  • Practice a pedigree for Valerie’s family


Points on gel electrophoresis l.jpg
Points on Gel Electrophoresis “Valerie”

  • 4-5 groups

  • Each group should pick up 5 samples labeled A, B, C, D and E

  • Load 20 ul

    • A: Ladder

    • B: wild type DNA “gold standard” (non-patient DNA)

    • C: Patient Peripheral Blood DNA

    • D Patient Tumor DNA

    • E Patient Non-tumor Breast Tissue


Gel electrophoresis l.jpg
Gel Electrophoresis “Valerie”

  • Heat samples 2 min. at 65 C in water bath in floating racks

  • Add 20 ul per well

  • Run for 75-125 volts, 20-25 minutes

  • Photograph results


What you hand in l.jpg
What you hand in: “Valerie”

  • Photograph with a legend and very brief explanation of the result

  • Family pedigree

  • Sequence of children and identify:

    • Who has a mutation

    • Where the mutation is (simply highlight with marker)


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