This tool is designed to generate primers for quick-change missense mutations easily on google Colab. Below are the instructions for preparing the necessary input files.
- Example Filename:
Shaker_FL.fa - Content: Start with a header line beginning with
>, followed by your sequence description. Write the DNA sequence in standard nucleotide codes (A, C, G, T) on the following lines. Sequence must be in frame. - Note: If the first codon does not correspond with the first aminoacid, this can be indicated by modifying the
starting_residuevalue
- Example Filename:
mutations.csv. - Content: List mutations as comma-separated values. Each line should be in the format
[position],[amino acid], where[position]is the residue number and[amino acid]is the desired amino acid using the single-letter code. Other options migth include "*" for ochre stop codon, "-" for amber and "X" for random mutation.
- Example Filename:
codontableXlaevis.csv. - Content: Include a mapping of amino acids to their respective codons. The first column lists the amino acids (single-letter code), followed by columns listing the high frequency codons.
Execute the cells in the notebook in order, ensuring that each cell completes successfully before proceeding to the next.
- Filename:
primers.csv - Content: List of primers as comma-separated values. First column list the name of the primer (
position_aa_Fw), second column is the primer sequence, the third column is the calculated Tm.