File S1 - Characterization of Ferredoxin-Dependent Glutamine-Oxoglutarate Amidotransferase (Fd-GOGAT) Genes and Their Relationship with Grain Protein Content QTL in Wheat

Supporting figures and text. Figure S1. Chromosome mapping of the Fd-GOGAT genes on chromosomes 2A and 2B. Genome specific markers were amplified from durum cultivars and hexaploid wheat cv Chinese Spring genetic stocks. A) The A-genome specific marker amplified in cvs Svevo, Ciccio, and Chinese Spring and sets of Chinese Spring nulli-tetrasomic deletion lines for chromosome group 2. The 350 bp fragment was absent in the nulli-2A-tetra-2B line, as indicated by arrows and confirming the localization on chromosome 2A. B) The B-genome specific marker amplified in cvs, Ciccio, Svevo, Chinese Spring and sets of nullitetrasomic deletion lines for chromosome group 2. The 450 bp fragment was absent in the nulli-2B-tetra-2A line, as indicated by arrows and confirming the localization on chromosome 2B. Figure S2. Alignment of wheat FD-GOGAT genes. Alignment of wheat cv Chinese Spring A, B, and D genome Fd-GOGAT genes from the beginning of exon 2 through the stop codons. Exons are indicated by red brackets and exon number above the alignments. The blue line indicates the signal peptide/mature polypeptide boundary. The stop codons are boxed in red. Figure S3. Fd-GOGAT signal sequences. The signal sequences encoded by six plant Fd-GOGAT genes are aligned with Clustal V: Wheat (A genome; present report and GAJL01283868), Brachypodium (BRADI1G19080), Rice (Os07g46460), Maize (NM_001112223), Arabidopsis (CP002688), Soybean. (AK245357). The red vertical line indicates the exon 1/exon 2 junction. The blue line indicates the end of the signal peptide. Figure S4. Alignment of wheat Fd-GOGAT polypeptides. Alignment of the three hexaploid wheat Fd-GOGAT polypeptides from the A, B, and D genomes. Differences in amino acid sequence are highlighted in yellow. Figure S5. Phylogenetic tree of Fd- and NADH-GOGAT proteins from diverse species. A selection of available GOGAT amino acid sequences from diverse phyla through genera were aligned with Clustal W and a phylogenetic tree formed by nearest-neighbor analysis. The wheat Fd-GOGAT homoeologues are in blue and red for the NADH-GOGAT version. Annotated sequences from other organisms have the same color coding as wheat. Unannotated sequences are in black. Figure S6. Comparison of Fd-GOGAT and NADH-GOGAT amino acid sequences. The amino acid sequences of the mature Fd-GOGAT from the A-genome is compared to the mature NADH-GOGAT from the A-genome by dot blot. Matching criteria was 80 match in a 5 amino acid residue window. Text S1. Fasta file of the three cv Chinese Spring Fd-GOGAT genes. Text S2. Fasta file of the three cv Chinese Spring Fd-GOGAT mature proteins. (PDF)