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Chapter 16 : Detection of Specific DNA Sequences: Hybridization Analysis

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Detection of Specific DNA Sequences: Hybridization Analysis, Page 1 of 2

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Abstract:

The activities in this lesson plan use paper models to illustrate basic concepts of hybridization analysis. Hybridization is a technique that takes advantage of the specificity of DNA base pairing for the detection of specific DNA sequences in a mixed sample. It is one of the fundamental methods for analysis of DNA. The first activity (Fishing for DNA) requires students to be familiar with the structure of DNA and the base-pairing rules. The second and third activities assume that students are familiar with restriction enzymes and the process of electrophoresis. If one plans to use the subsequent lessons on DNA sequencing, the polymerase chain reaction, or the analysis of human DNA, it is important to introduce one`s class to hybridization. Restriction digestion, electrophoresis, and staining allow scientists to cut DNA molecules into reproducible fragments and to look at the sizes of those fragments. In brief, hybridization analysis involves separating the strands of (denaturing) the DNA molecules to be analyzed and then mixing those separated strands with many copies of a single-stranded DNA or RNA molecule. This single-stranded DNA or RNA molecule has the complement of the base sequence of interest and is labeled for detection (often with radioactive isotopes). It is called a probe.

Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16

Key Concept Ranking

Agarose Gel Electrophoresis
0.43467104
DNA Restriction Enzymes
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Figures

Image of Figure 16.1
Figure 16.1

Colony hybridization

Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16
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Image of Figure 16.2
Figure 16.2

Assembling a Southern blot. (A) The student is placing the membrane on top of the gel. (B) The complete transfer setup. The bottle on top is simply a weight.

Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16
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Image of Figure 16.3
Figure 16.3

Southern hybridization analysis. (A) Diagrammatic representation. (B) Two stained gel lanes and the results of a hybridization to those lanes. kb, DNA fragment size in kilobase pairs.

Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16
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Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16
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Image of Diagram 16.I
Diagram 16.I

Hybridization of a probe to a sequence within single-stranded sample DNA.

Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16
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Image of Diagram16.II
Diagram16.II

Southern hybridization analysis shows which restriction fragments hybridize to a probe.

Citation: Kreuzer H, Massey A. 2008. Detection of Specific DNA Sequences: Hybridization Analysis, p 257-269. In Molecular Biology and Biotechnology: A Guide for Teachers, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816100.ch16
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References

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