BIOB 160 Principles of Living Systems · Unit 4: Molecular genetics · on Final exam
4.5 DNA technology
Study notes for DNA technology in BIOB 160 Principles of Living Systems at Montana State University: the big ideas, key formulas, a worked example, common mistakes and an exam tip. The same topic has endless practice questions and flashcards in Mathub.
Big ideas
- Restriction enzymes cut DNA at specific sequences (EcoRI cuts GAATTC), often leaving sticky ends. Fragments with matching ends are joined by DNA ligase to make recombinant DNA.
- Gene cloning: insert a gene into a plasmid vector, transform bacteria, select with an antibiotic-resistance gene, and let the bacteria copy it.
- Gel electrophoresis separates DNA fragments by size: DNA is negatively charged and moves toward the positive electrode, and smaller fragments move farther.
- PCR copies a target sequence: denature at about 95 °C, anneal primers at about 50–65 °C, extend with heat-stable Taq polymerase at about 72 °C. Each cycle doubles the copies: 2ⁿ after n cycles.
- Sanger sequencing uses dideoxynucleotides that stop synthesis; next-generation methods sequence millions of fragments at once. CRISPR-Cas9 uses a guide RNA to lead the Cas9 nuclease to a target sequence, where it cuts both strands so the gene can be disrupted or edited.
Key formulas
PCR copies
$$N = N_0 \cdot 2^{n}$$
Worked example
You start PCR with 10 copies of a target. How many copies after 20 cycles, assuming perfect doubling?
Show the solution
N = 10 × 2²⁰ = 10 × 1,048,576 ≈ 10.5 million copies.
Common mistakes
- Saying DNA moves toward the negative electrode. It is negatively charged, so it moves toward the positive one.
- Thinking larger fragments travel farther. Smaller ones do.
- Forgetting why Taq polymerase is used: it survives the denaturing temperature.
Exam tipReading: Campbell ch. 19 (pp. 413–420), per the syllabus. For PCR arithmetic, use 2ⁿ and powers of ten (2¹⁰ ≈ 1,000).
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