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How can I assemble long, single-stranded oligos to create several kb, clonable gene sequences (assembly PCR)?
Assembly PCR is a very flexible technique for producing novel gene sequences from single-stranded oligos or a mix of single- and double-stranded DNA. Several single-stranded oligonucleotides, such as the IDT 60–120 nt Ultramer® Oligonucleotides, can be designed with short overlapping sequences and then assembled in a 2-step process to produce longer genes of up to several thousand base pairs. Considerations for setting up such reactions and a brief protocol can be found in the article, Assembly PCR for Novel Gene Synthesis, in DECODED 2.4, October 2012 (www.idtdna.com/decoded). IDT also offers gBlocks® Gene Fragments, custom synthesized, double-stranded DNA fragments that can streamline the assembly and cloning process. Learn more at www.idtdna.com/gblocks.