- Course Introduction
- 01: Evidence & DNA
- 02: Forensic Biology
- 03: DNA Extraction & Quantitation
- 04: DNA Amplification
- Overview
- Introduction
- Amplification
- Primer Design
- Primer Length
- Primer Melting Temperature
- Primer Annealing Temperature
- Values for Tm
- Maximum 3′ Stability
- GC Content Percentage
- No Secondary Structures
- No Self-complementarity
- No Complementarity To Other Primers
- No Long Runs With The Same Base
- Distance Between Two Primers On Target Sequence
- Plateau Effect
- Thermal Cyclers
- Thermal Cycling Parameter and Optimization
- Works Cited & Online Links
- Locus Selection
- Multiplexing
- Contamination
- Overview
- 05: Amplified DNA Product Separation
- 06: STR Data Analysis & Interpretation
- 07: Population Genetics & Statistics
- 08: Communicating Results
- 09: Other DNA Markers & Technologies
- Course Resources
- Laboratory Training Manual
President's DNA Initiative
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Advancing Justice Through DNA Technology
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Advancing Justice Through DNA Technology
DNA Analyst Training
Buffer and MG2+
Home > DNA Amplification > Overview > Thermal Cyclers >Buffer and MG2+
Buffer
Reaction pH can alter the fidelity of an insertion. A 10-50µM Tris-HCl buffer with a pH between 8.3 and 8.8 (when measured at 20°C) is used.11
Mg2+
The magnesium concentration may affect all of the following:11
- primer annealing
- strand disassociation temperatures of template
- strand disassociation temperatures of PCR product
- product specificity
- formation of primer-dimer artifacts
- enzyme activity and fidelity
In general, increasing the free magnesium concentration increases yield and decreases specificity and fidelity.
Taq DNA polymerase requires free magnesium (0.5 to 2.5mM) additional to that bound by template DNA, primers, and dNTPs. The presence of EDTA or other chelators in the primer stock or template DNA might disturb the apparent magnesium optimum.
