Biotechnology

Biotechnology cuts DNA with restriction enzymes, joins pieces with ligase, and amplifies targets with PCR. Restriction enzymes recognize palindromic sequences and leave sticky ends, and each PCR cycle of denaturation, annealing, and extension doubles the DNA. Putting a gene into a vector and expressing it in a host can mass-produce substances such as insulin. Slide cut progress and PCR cycle count to see how amplification grows.

Intro — Genes can be 'edited'

💡 Core tools of biotechnology

①Restriction enzymes: 'molecular scissors' that cut DNA at specific sites

②PCR: 'molecular copier' that amplifies DNA fragments millions of times

③Ligase: 'molecular glue' that joins cut DNA

④COVID PCR test = amplifies viral DNA for detection

⑤Recombinant DNA tech mass-produces insulin, growth hormones, etc.

Restriction Enzymes — Molecular Scissors

Cleavage progress0

🔍 How restriction enzymes work

①Palindrome recognition: same sequence read top-down or bottom-up

②EcoRI: 5'-G|AATTC-3' / 3'-CTTAA|G-5'

③Cuts each strand at different positions → sticky ends

④Sticky ends pair complementarily → joins different DNA fragments

⑤Blunt-end enzymes also exist (cut both strands at same position)

PCR — Gene Amplification

PCR cycles10

PCR amplification

DNA amount = 2ⁿ (n = cycles)

30 cycles → ~1 billion copies!

💡 Remember the 3 PCR steps

①Denaturation (94°C): heat separates double-strand DNA

②Annealing (55°C): primers bind specific sites on each strand

③Extension (72°C): heat-stable DNA polymerase (Taq) synthesizes new strand

④These 3 steps = 1 cycle, typically repeated 25–35 times

⑤COVID PCR test: viral RNA → cDNA → PCR amplification → fluorescent detection

Recombinant DNA and Applications

Recombinant DNA process

restriction cut → vector insertion (ligase) → host transformation

introduce target gene into another organism for expression

📐 Key techniques and applications

①Vector: typically plasmid — self-replicates in bacteria

②Transformation: bacteria with foreign gene express it

③Mass insulin production: human insulin gene inserted into E. coli

④Gel electrophoresis: separates DNA by size (smaller moves faster)

⑤DNA fingerprinting: STR repeat sequences differ between individuals

Worked Examples and Exam Practice

Example 1

If you amplify 1 DNA molecule by PCR for 10 cycles, how many copies result in theory? (doubling each cycle)

PCR doubles the DNA each cycle, so after n cycles there are 2ⁿ times as many.

For 10 cycles, 2¹⁰ = 1024, so 1 molecule becomes about 1024.

▸ 2¹⁰ = 1024 copies

Exponential amplification explodes with more cycles: about 1 million at 20 cycles, about 1 billion at 30.

Example 2

After cutting two different DNAs with the same restriction enzyme to make sticky ends, which enzyme joins the two pieces into one?

Cutting with the same enzyme gives both pieces complementary sticky-end sequences.

Once the ends pair up, DNA ligase seals the sugar-phosphate backbone into one recombinant DNA.

▸ DNA ligase

A restriction enzyme is the scissors, ligase is the glue. This pair inserts a target gene into a vector such as a plasmid.

School-exam style

Which statement about biotechnology is correct?

①The extension step of PCR is about 94°C

②Restriction enzymes cut DNA at random sites

③After n cycles of PCR, DNA is amplified 2ⁿ times

④In gel electrophoresis, larger DNA fragments move faster

⑤DNA ligase is an enzyme that cuts DNA

▸ ③ After n cycles of PCR, DNA is amplified 2ⁿ times

PCR repeats denaturation, annealing, and extension as one cycle; the DNA doubles each cycle, giving 2ⁿ after n cycles.

Extension is about 72°C (①), restriction enzymes cut specific palindromes (②), smaller fragments move faster in electrophoresis (④), and ligase joins DNA (⑤).

Summary

PCR amplification

2ⁿ copies (n = cycles)

20 cycles ≈ 1M, 30 cycles ≈ 1B copies

🎯 Exam Points

①PCR 3 steps: denaturation (94°C) → annealing (55°C) → extension (72°C)

②n cycles → 2^n DNA (exponential amplification)

③Restriction enzymes: palindrome recognition, sticky/blunt ends

④Recombinant DNA: enzymes + ligase + vector (plasmid)

⑤Applications: GMO, gene therapy, stem cells, DNA fingerprinting, PCR test

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Gene Regulation

Biodiversity & Classification

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