Example Data Table
| Sequence A | Sequence B | Match | Mismatch | Gap Open | Gap Extend | Score | Identity |
|---|---|---|---|---|---|---|---|
ACGTTGCA |
A-GTCGCA |
2 | -1 | -2 | -1 | 9 | 75% |
MKT-ALIV |
MRTQAL-V |
BLOSUM62 | Matrix-based | -5 | -1 | Matrix dependent | Varies |
Formula Used
General alignment score
Total Score = Σ substitution scores + Σ gap opening penalties + Σ gap extension penalties
Simple nucleotide scoring
Score(i) = Match Score when the residues are equal.
Score(i) = Mismatch Penalty when the residues differ.
Transition or transversion mode
Transitions are purine-to-purine or pyrimidine-to-pyrimidine substitutions. Transversions switch residue classes and often receive a stronger penalty.
Gap handling
Gap Block Score = Gap Open + (Gap Length - 1) × Gap Extend
Normalized score
Normalized Score = Raw Score ÷ Alignment Length
Identity percentage
Identity % = (Matches ÷ Alignment Length) × 100
How to Use This Calculator
- Paste two already aligned biological sequences into the input boxes.
- Select DNA, RNA, protein, or custom mode.
- Choose a nucleotide model or protein substitution matrix.
- Set match, mismatch, gap opening, and gap extension values.
- Decide whether terminal gaps or double-gap columns should affect scoring.
- Press Calculate Alignment Score to view the summary and detailed position table.
- Use the CSV or PDF buttons after calculation to export your results.
Frequently Asked Questions
1. What does this calculator measure?
It measures the score of an alignment you already have. The score reflects matches, mismatches, substitutions, and gap penalties across every aligned position.
2. Does this tool create the alignment for me?
No. It evaluates an existing alignment. Use a separate alignment algorithm if you need to generate the best path before scoring it here.
3. When should I use transition and transversion scoring?
Use it for DNA or RNA when you want biologically meaningful penalties. Transitions often occur more frequently than transversions, so researchers may score them differently.
4. Why would I ignore terminal gaps?
Semi-global comparisons often ignore leading or trailing gaps. This helps when one sequence is shorter or when incomplete ends should not reduce the score.
5. What are positive substitutions in protein mode?
They are non-identical residue pairs that still receive a positive matrix score. This suggests conservative replacement between amino acids with similar biochemical properties.
6. Why must the sequences have equal length?
Each column represents one comparison event. Unequal lengths mean the alignment is incomplete, so the calculator cannot correctly score every position.
7. Can I use ambiguous symbols like N or X?
Yes. The calculator treats common ambiguous symbols as wildcard positions and scores them using the wildcard value you provide.
8. What is the difference between raw and normalized score?
Raw score is the full total across the alignment. Normalized score divides that value by alignment length, which makes comparisons easier across different alignment sizes.