I'm trying to design a primer for Gibson assembly. My gene of interest is on a plasmid, and I want to copy that gene, and put it into a different plasmid. I am unsure how to design my primers for PCR. I know that I need 2 sets of primers (4 total). 1. Backward and forward primer to copy my gene out of the plasmid that holds it. 2. Backward and forward primer to copy the plasmid. I also know that I need to complementary regions of overlap between my gene and the plasmid i want to put it in. How do I do this? Here is the Plasmid and the gene. I want to place the gene in between the biobrick prefix and suffix. Here is what primers I think I need: For the plasmid I want to put the gene in, my primers should be - Forward: ATTCGCGGCCGCTTCTAGAG - Reverse: CTGACGCGGCCGCTACTAGTA Primers to copy gene and add overlap - Reverse:ATCATTTCAAATTCGTCCATCTCTAGAAGCGGCCGCGAAT - Forward: AATTAGAAAGAGATTATAATACTAGTAGCGGCCGCTGCA Here is the gene - [IodoY TyrRs][1] [Here is the plasmid][2] I want to put it in. I am using geneious, but I think for this purpose, word works just fine. [1]: https://docs.google.com/file/d/0B5gGZ4-SD7-ZalhLdG5PR3VvUVk/edit?usp=sharing [2]: https://docs.google.com/file/d/0B5gGZ4-SD7-ZbUNld2FWel9SQTA/edit?usp=sharing