From 2007.igem.org

Beta portion pcr CA1154F/G00101 on 9145-I716072 (575 bp) Product is ADtemp1 Sub ADtemp1 into pTRC plasmid with (Bgl2/XhoI) Product is ADtemp2 pcr CA1099F/D58 on ADtemp2 (3465 bp) Product is ADtemp3 _______________________________________________________ Alpha portion Pcr CA1150F/CA1099R on I716073 (1727 bp) Product is ADtemp5 ____________________________________________________________ Digest ADtemp3 (BsaI/BamHI) Digest ADtemp5 (BsaI/BglII) Ligate, product is: I716074 (Best yield)