Transgene Construction

For truncation constructs, pHGW (Karimi et al., 2002) digested with EcoRV and self-ligated was used as a binary vector. A NOS terminator was inserted between the SacI and HindIII sites of this vector. All sequences, including the SLAC1 promoter (−1568 to +1) and GFP, were isolated by PCR using the previously described pBI101-SLAC1-GFP construct (Negi et al., 2008) and inserted between the SalI and SacI sites of the binary vector. For ΔN, ΔC, and ΔNC, genomic sequences corresponding to the coding regions for amino acids 180 to 556, 1 to 507, and 180 to 507 of the SLAC1 protein were used, respectively. All primers used for plasmid construction are listed in Supplemental Table 1. For single amino acid substitution constructs, a fragment including the single amino acid substitution was prepared by recombinant PCR and inserted into the corresponding sites of pBI101-SLAC1-GFP (Negi et al., 2008). The DNA sequences harboring mutations causing the amino acid substitutions are listed in Supplemental Table 2.