PCR Protocol - Biology Process

Description

Polymerase chain reaction: thermostable-polymerase amplification of a target DNA sequence through repeated denaturation, primer annealing, and extension, with a gel size check before the product is accepted.

Process Flowchart

graph TD N1["Template DNA"] N2["Forward + reverse primers"] N3["dNTPs, Taq, Mg2+ buffer"] N4["Assemble mix on ice"] N5["Initial denaturation 95C"] N6["Denature 94C"] N7["Anneal primers ~55C"] N8["Extend at 72C (Taq)"] N9{"Cycles complete?"} N10["Final extension 72C"] N11{"Correct amplicon size?"} N12["Amplified target DNA"] N1 --> N4 N2 --> N4 N3 --> N4 N4 --> N5 N5 --> N6 N6 --> N7 N7 --> N8 N8 --> N9 N9 -->|no| N6 N9 -->|yes| N10 N10 --> N11 N11 -->|optimize| N4 N11 -->|yes| N12 style N1 fill:#ff6b6b,color:#fff style N2 fill:#ff6b6b,color:#fff style N3 fill:#ffd43b,color:#000 style N4 fill:#51cf66,color:#fff style N5 fill:#51cf66,color:#fff style N6 fill:#51cf66,color:#fff style N7 fill:#51cf66,color:#fff style N8 fill:#51cf66,color:#fff style N9 fill:#74c0fc,color:#fff style N10 fill:#51cf66,color:#fff style N11 fill:#74c0fc,color:#fff style N12 fill:#b197fc,color:#fff

Node provenance

Each step below is grounded in a cited source (hover a citation for the full title).

Step	Role	Source
Template DNA	Triggers & Inputs	Saiki, R. K. et al. 1988
Forward + reverse primers	Triggers & Inputs	Mullis, K. B. et al. 1987
dNTPs, Taq, Mg2+ buffer	Structures & Objects	Saiki, R. K. et al. 1988
Assemble mix on ice	Processing & Operations	Sambrook, J. et al. 2001
Initial denaturation 95C	Processing & Operations	Saiki, R. K. et al. 1988
Denature 94C	Processing & Operations	Mullis, K. B. et al. 1987
Anneal primers ~55C	Processing & Operations	Mullis, K. B. et al. 1987
Extend at 72C (Taq)	Processing & Operations	Saiki, R. K. et al. 1988
Cycles complete?	Intermediates & States	Mullis, K. B. et al. 1987
Final extension 72C	Processing & Operations	Sambrook, J. et al. 2001
Correct amplicon size?	Intermediates & States	Sambrook, J. et al. 2001
Amplified target DNA	Products & Outputs	Saiki, R. K. et al. 1988

🎨 Color Scheme (5-Color System)

Red
Triggers & Inputs

Yellow
Structures & Objects

Green
Processing & Operations

Blue
Intermediates & States

Violet
Products & Outputs

Process Statistics

Nodes: 12
Edges: 13
Conditionals: 2
AND Gates: 2
OR Gates: 2
Total Gates: 4

Keywords

pcr
amplification
taq polymerase
thermal cycling
primers

Sources & References

Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase
Saiki, R. K.; Gelfand, D. H.; Stoffel, S.; Scharf, S. J.; Higuchi, R.; Horn, G. T.; Mullis, K. B.; Erlich, H. A. , Science (1988)
DOI: 10.1126/science.2448875
Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction
Mullis, K. B.; Faloona, F. A. , Methods in Enzymology (1987)
DOI: 10.1016/0076-6879(87)55023-6
Molecular Cloning: A Laboratory Manual (3rd ed.)
Sambrook, J.; Russell, D. W. , Cold Spring Harbor Laboratory Press (2001)
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