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Primer Reconstitution Formula:
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Primer reconstitution is the process of dissolving lyophilized primers in an appropriate volume of solvent (usually TE buffer or nuclease-free water) to achieve a desired concentration for molecular biology experiments.
The calculator uses the primer reconstitution formula:
Where:
Explanation: The formula converts nmol to pmol (×1000) and divides by the desired concentration in μM to calculate the volume in μL needed to achieve that concentration.
Details: Accurate primer reconstitution is essential for consistent experimental results in PCR, sequencing, and other molecular biology techniques. Incorrect concentrations can lead to failed reactions or suboptimal results.
Tips: Enter the amount of primer (as provided by the manufacturer, typically in nmol) and your desired working concentration (typically 100 μM for stock solutions). The calculator will determine the volume of solvent needed.
Q1: What is a typical stock concentration for primers?
A: Most researchers prepare 100 μM stock solutions, which can then be diluted further for working solutions (typically 10 μM).
Q2: What solvent should I use for reconstitution?
A: TE buffer (10 mM Tris, 1 mM EDTA, pH 8.0) or nuclease-free water are most common. TE provides better stability for long-term storage.
Q3: How should I store reconstituted primers?
A: Store at -20°C in small aliquots to avoid repeated freeze-thaw cycles. Stock solutions are typically stable for years when properly stored.
Q4: What if my primer amount is in μg instead of nmol?
A: You'll need to convert μg to nmol using the molecular weight of the primer. The formula is: nmol = (μg × 1000) / (MW in g/mol).
Q5: How do I calculate the molecular weight of my primer?
A: MW can be calculated as: (A × 313.2) + (T × 304.2) + (C × 289.2) + (G × 329.2) - 61.0, where A/T/C/G are the counts of each nucleotide.