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会计学
PCR与亲子刑事鉴定
2021/4/25
DNA template (double stranded)
primers (single stranded DNA)
DNA polymerase
nucleotides (A,C,G,T)
thermocycler
PCR: Requirements
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Thermocycler
- controlled temperature
- controlled rate of temperature change
PCR: Requirements
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Step 1: Separate the two strands of DNA template
usually around 95 C
Step 2: Cool to let the primers bind to the template strands
usually around 40 to 60 C
Step 3: DNA polymerase extends the primers
two new complementary DNA strands synthesized
usually around 72 C
PCR: 3 steps
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Taq polymerase
Need heat-stable DNA polymerase: Taq polymerase
from a thermophilic (heat-loving) bacteria
not inactivated by repeated heat treatments of PCR step 1
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Cycles of PCR:
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◎ Avoid contamination
◎ Component
dH2O
10 X PCR buffer
dNTP
sense primer
antisense primer
Taq DNA polymerase
Polymerase Chain Reaction, PCR
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◎ Negative control / Positive control / Exp.
◎ Detect PCR products
EtBr stained agarose gel electrophoresis
Southern blot
◎ Optimization
enzyme
dNTP
Mg2+
Temp. of annealing, extension and denaturation
cycle no.
primers
Polymerase Chain Reaction, PCR
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