How do I split the 15-nt homologous overlap among adjacent DNA fragments?

The homologous 15-nt overlap can be split between two adjacent DNA fragments. It can only be split between an insert and vector if the vector is linearized via inverse PCR. Primer design for this option is not facilitated by the online Primer Design Tool. We recommend SnapGene Viewer as a helpful, free online tool to help with this task.In-Fusion Cloning [...]

Can I use In-Fusion to insert external nucleotide sequences?

Small external nucleotide sequences (e.g., small tags, Kozak sequences, restriction sites, cleavage sites, etc.) can be added between the template/gene-specific portion and the 15-nt homologous overlap of the In-Fusion PCR primer.In-Fusion Cloning [...]

Can I modify translational reading frames during In-Fusion?

In-Fusion Cloning makes it possible to seamlessly clone your gene of interest in the same translational reading frame as a desired tag (e.g., fluorescent protein, Myc, HA, etc.). Specifics in the design of In-Fusion PCR primers facilitate this application, allowing you to maintain reading frame continuity in the recombinant vector. Two options for doing so are below: -Adjusting the length of [...]

How do I preserve restriction-site using In-Fusion?

Primer design lets you easily preserve or eliminate the restriction sites used to linearize the cloning vector. In order to maintain restriction sites at cloning junctions, nucleotides can be added to the PCR primers between the template-specific portion and the 15-nt homologous overlap. The online Primer Design Tool offers the option to preserve the restriction site(s) (the Primer Design Tool [...]

What are the features of Takara online primer design tools?

Instructions for designing In?Fusion PCR primers are included in all In?Fusion Cloning user manuals. Several tools are also available to help with the process. 1. Our online Primer Design Tool facilitates primer design for In?Fusion Cloning, and is compatible with Mozilla Firefox or Google Chrome web browsers. (Internet Explorer is not compatible with the Primer Design Tool.) Specific cloning [...]