Gibson Assembly® Site Directed Mutagenesis (SDM) Primer Design Tool

PLEASE READ BEFORE BEGINNING

Gibson Assembly SDM primer tool is developed to design primers to incorporate single or multiple substitutions, insertion or deletions using Gibson Assembly Site Directed Mutagenesis kit (Cat. No. GA2100-S, GA 2100-10). To use this tool, user will input sequences and define the mutations to be incorporated by PCR reaction. The tool designs primers that introduce mutation and create homologous sequence at the end of adjacent fragments. The homologous sequences allow for scarless DNA assembly using the GA SDM Assemble mix.

The SGI-DNA Gibson Assembly® SDM Primer Tool can be used in the following browsers:

  • The Google Chrome™ browser
  • Firefox
  • Internet Explorer 10 (IE10)

For optimal results, use Google Chrome.

For more information about the Gibson Assembly® method visit sgidna.com

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1. Enter Your Sequence

The Gibson Assmbly SDM Primer Design Tool assumes the input sequence to be a linear template. The first nucleotide will be numerically indexed as 1.

Enter Your Vector Sequence Below (FASTA or plain text)

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2. Mutation Input

To design Insertion mutation, click and drag to highlight (or directly enter) the location of the two nucleotides between which the insertion will take place. Click the blue + box to open a new mutation definition line. Select “INSERTION” from the dropdown menu. Then enter the insertion sequence in the Replacement String box.

To design a deletion mutation, click and drag to highlight (or directly enter) the location of the region to be deleted. Select “DELETION” from the dropdown menu, and leave the Replacement String box blank.

To design a substitution mutation, click and drag to highlight (or directly enter) the location of the region to be replaced. Select “SUBSTITUTION” from the dropdown menu, and enter the substitution sequence in the Replacement String box. Click save to confirm mutation design.

Make up to 5 edits

Reading Frame:




Type Location (base pair) Replacement String
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3. Confirm Mutations



Pairwise Alignment
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4. Primer information and assembly graphic

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